The germination of pollen grain in vitro and the growth of pollen tube in the pistil of rice were observed with a microscope. The stigma was removed at different time points after pollination to study its effect on se...The germination of pollen grain in vitro and the growth of pollen tube in the pistil of rice were observed with a microscope. The stigma was removed at different time points after pollination to study its effect on seed setting rate. The rice pollen grain started to germinate at 2 min after pollination and the pollen tube penetrated stigma into style in 5-10 min, 30 min later the end of pollen tube reached the bottom of ovary, and only some pollen tubes arrived at embryo sac at 40 min after pollination. Meanwhile, a small amount of callose began to deposit in the pollen tubes, a great deal of callose was observed at 50 min after pollination, whereas the pollen grain began to shrink. The growing rates of pollen tube in the rice stigma, style and ovary were 1500, 5000, and 5400 μm/h, respectively. The seed setting rate was quite low when the stigma was removed at about 10-15 min after pollination, gradually increased when it removed at 20 min to 50 min after pollination, and over 60% when it removed at 50 min after pollination and finally tended to be stable.展开更多
Pollen tube growth is essential for successful double fertilization, which is critical for grain yield in crop plants. Rapid alkalinization factors(RALFs) function as ligands for signal transduction during fertilizati...Pollen tube growth is essential for successful double fertilization, which is critical for grain yield in crop plants. Rapid alkalinization factors(RALFs) function as ligands for signal transduction during fertilization. However, functional studies on RALF in monocot plants are lacking.Herein, we functionally characterized two pollen-specific RALFs in rice(Oryza sativa) using multiple clustered regularly interspaced palindromic repeats(CRISPR)/CRISPR-associated protein 9-induced loss-of-function mutants,peptide treatment, expression analyses, and tag reporter lines. Among the 41 RALF members in rice, OsRALF17 was specifically expressed at the highest level in pollen and pollen tubes.Exogenously applied OsRALF17 or OsRALF19peptide inhibited pollen tube germination and elongation at high concentrations but enhanced tube elongation at low concentrations, indicating growth regulation. Double mutants of OsRALF17 and OsRALF19(ralf17/19) exhibited almost full male sterility with defects in pollen hydration, germination, and tube elongation,which was partially recovered by exogenous treatment with OsRALF17 peptide. This study revealed that two partially functionally redundant OsRALF17 and OsRALF19 bind to Oryza sativa male-gene transfer defective 2(OsMTD2) and transmit reactive oxygen species signals for pollen tube germination and integrity maintenance in rice. Transcriptomic analysis confirmed their common downstream genes, in osmtd2 and ralf17/19. This study provides new insights into the role of RALF, expanding our knowledge of the biological role of RALF in regulating rice fertilization.展开更多
Due to self-incompatibility(SI)prevents self-fertilization,natural or artificial cross-pollination has been conducted in many orchards to stabilize fruit yield.However,it is still puzzled which routes of self S-RNase ...Due to self-incompatibility(SI)prevents self-fertilization,natural or artificial cross-pollination has been conducted in many orchards to stabilize fruit yield.However,it is still puzzled which routes of self S-RNase arresting pollen tube growth.Herein,17 COBRA genes were isolated from pear genome.Of these genes,the pollen-specifically expressed PbCOB.A.1 and PbCOB.A.2 positively mediates pollen tube growth.The promoters of PbCOB.A.1 and/or PbCOB.A.2 were bound and activated by PbABF.E.2(an ABRE-binding factor)and PbC2H2.K16.2(a C2H2-type zinc finger protein).Notably,the expressions of PbCOB.A.1,PbCOB.A.2,and PbC2H2.K16.2 were repressed by self S-RNase,suggesting that self S-RNase reduces the expression of PbCOB.A.1 and PbCOB.A.2 by decreasing the expression of their upstream factors,such as PbC2H2.K16.2,to arrest pollen tube growth.PbCOB.A.1 or PbCOB.A.2 accelerates the growth of pollen tubes treated by self S-RNase,but can hardly affect level of reactive oxygen species and deploymerization of actin cytoskeleton in pollen tubes and cannot physically interact with any reported proteins involved in SI.These results indicate that PbCOB.A.1 and PbCOB.A.2 may not relieve S-RNase toxicity in incompatible pollen tube.The information provides a new route to elucidate the arresting pollen tube growth during SI reaction.展开更多
With radical global climate change and global warming, high temperature stress has become one of major factors exerting a major Influence on crop production, In the cotton (Gossyplum hirsutum L.)-growlng areas of Ch...With radical global climate change and global warming, high temperature stress has become one of major factors exerting a major Influence on crop production, In the cotton (Gossyplum hirsutum L.)-growlng areas of China, especially in the Yangtze River valley, unexpected periodic episodes of extreme heat stress usually occur In July and August, the peak time of cotton flowering and boll loading, resulting In lower boll set and lint yield. Breeding programs for screening high temperature-tolerant cotton germplasm and cultlvars are urgent In order to stabilize yield in the current and future warmer weather conditions. In the present study, 14 cotton cultivars were quantified for in vitro pollen germination and pollen tube growth in response to temperatures ranging from 10 to 50 ℃ at 5 ℃ intervals. Different cotton genotypes varied In their in vitro pollen germination and pollen tube length responses to the different temperatures. Maximum pollen germination and pollen tube length ranged from 25.2% to 56.2% and from 414 to 682 pro, respectively. The average cardinal temperatures (Tmin,, Topt, and Tmax) also varied among the 14 cultivars and were 11.8, 27.3, and 42.7 ℃ for pollen germination and 11.8, 27.8, and 44.1 ℃ for maximum pollen tube length. Variations In boll retention and boll numbers per plant in field experiments were found for the 14 cotton cultivars and the boll retention and boll retained per plant on 20 August varied considerably In different years according to weather conditions. Boll retention on 20 August was highly correlated with maximum pollen germination (R^2 = 0.84) and pollen tube length (R^2=0.64). A screening method based on principle component analysis of the combination of pollen characterlatics In an in vitro experiment and boll retention testing In the field environment was used In the present study and, as a result, the 14 cotton cultlvars could be classified as tolerant, moderately tolerant, moderately susceptible and susceptible to high temperature.展开更多
Ca2+ has been established as an important second messenger regulating pollen germination and tube growth. However, to date, only a few signaling components have been identified to decode and relay Ca2+ signals in gr...Ca2+ has been established as an important second messenger regulating pollen germination and tube growth. However, to date, only a few signaling components have been identified to decode and relay Ca2+ signals in growing pollen tubes. Here, we report a function for the calcineurin B-like (CBL) Ca2+ sensor proteins CBL1 and CBL9 from Arabidopsis in pollen germination and tube growth. Both proteins are expressed in mature pollen and pollen tubes and impair pollen tube growth and morphology if transiently overexpressed in tobacco pollen. The induction of these phenotypes requires efficient plasma membrane targeting of CBL1 and is independent of Ca2+ binding to the fourth EF-hand of CBL1. Overexpression of CBL1 or its closest homolog CBL9 in Arabidopsis renders pollen germination and tube growth hypersensitive towards high external K+ concentrations while disruption of CBL1 and CBL9 reduces pollen tube growth under low K~ conditions. Together, our data identify a crucial function for CBL1 and CBL9 in pollen germination and tube growth and suggest a model in which both proteins act at the plasma membrane through regulation of K+ homeostasis.展开更多
Dynamics of the actin cytoskeleton are essential for pollen germination and pollen tube growth. ACTIN- DEPOLYMERIZlNG FACTORs (ADFs) typically contribute to actin turnover by severing/depolymerizing actin filaments....Dynamics of the actin cytoskeleton are essential for pollen germination and pollen tube growth. ACTIN- DEPOLYMERIZlNG FACTORs (ADFs) typically contribute to actin turnover by severing/depolymerizing actin filaments. Recently, we demonstrated that Arabidopsis subclass III ADFs (ADF5 and ADF9) evolved F-actin-bundling function from conserved F-actin-depolymerizing function. However, little is known about the physiological function, the evolutional significance, and the actin-bundling mechanism of these neo- functionalized ADFs. Here, we report that loss of ADF5 function caused delayed pollen germination, retarded pollen tube growth, and increased sensitive to latrunculin B (LatB) treatment by affecting the generation and maintenance of actin bundles. Examination of actin filament dynamics in living cells revealed that the bundling frequency was significantly decreased in adf5 pollen tubes, consistent with its biochem- ical functions. Further biochemical and genetic complementation analyses demonstrated that both the N- and C-terminal actin-binding domains of ADF5 are required for its physiological and biochemical functions. Interestingly, while both are atypical actin-bundling ADFs, ADF5, but not ADF9, plays an important role in mature pollen physiological activities. Taken together, our results suggest that ADF5 has evolved the function of bundling actin filaments and plays an important role in the formation, organization, and maintenance of actin bundles during pollen germination and pollen tube growth.展开更多
The present study is expected to provide better understanding of how exo-β-glucanase acts on the biochemo-rheological property of the cell wall, and thus the regulation mechanism of pollen tube growth. A polymerase c...The present study is expected to provide better understanding of how exo-β-glucanase acts on the biochemo-rheological property of the cell wall, and thus the regulation mechanism of pollen tube growth. A polymerase chain reaction (PCR)-based strategy and cDNA library screening were used to clone an exo-β-glucanase gene (LP-Exol) from Lilium Iongiflorum pollen tubes. The LP-Exol sequence showed a high level of homology to that of reported plant exo-β-glucan hydrolases. Phylogenetic analysis demonstrated that there was close relationship between LP-Exol and other exo-glucan hydrolases in Poacea plants. Northern analysis indicated that LP-Exol transcripts stored in pollen grains at low levels; LP-Exol transcription level enhanced at 2 h after the onset of pollen incubation and remained relatively high during pollen tube elongation. Moreover, the expression of LP-Exol was undetectable in petals, filaments, stigmas, styles, ovaries, leaves, and stems. These results suggest that LP-Exol gene is a late pollen gene, specifically contributing to the regulation of lily pollen germination and pollen tube growth.展开更多
Although pollen tube growth is a prerequisite for higher plant fertilization and seed production, the processes leading to pollen tube emission and elongation are crucial for understanding the basic mechanisms of tip ...Although pollen tube growth is a prerequisite for higher plant fertilization and seed production, the processes leading to pollen tube emission and elongation are crucial for understanding the basic mechanisms of tip growth. It was generally accepted that pollen tube elongation occurs by accumulation and fusion of Golgi-derived secretory vesicles (SVs) in the apical region, or clear zone, where they were thought to fuse with a restricted area of the apical plasma membrane (PM), defining the apical growth domain. Fusion of SVs at the tip reverses outside cell wall material and provides new segments of PM. However, electron microscopy studies have clearly shown that the PM incorporated at the tip greatly exceeds elongation and a mechanism of PM retrieval was already postulated in the mid-nineteenth century. Recent studies on endocytosis during pollen tube growth showed that different endocytic pathways occurred in distinct zones of the tube, including the apex, and led to a new hypothesis to explain vesicle accumulation at the tip; namely, that endocytic vesicles contribute substantially to V-shaped vesicle accumulation in addition to SVs and that exocytosis does not involve the entire apical domain. New insights suggested the intriguing hypothesis that modulation between exo- and endocytosis in the apex contributes to maintain PM polarity in terms of lipid/protein composition and showed distinct degradation pathways that could have different functions in the physiology of the cell. Pollen tube growth in vivo is closely regulated by interaction with style molecules. The study of endocytosis and membrane recycling in pollen tubes opens new perspectives to studying pollen tube-style interactions in vivo.展开更多
[Objective] This study aimed to investigate the influence of different plant growth regulators on apricot pollen germination and pollen tube growth. [Method] Pollens of six kinds of Xinjiang apricots were cultured in ...[Objective] This study aimed to investigate the influence of different plant growth regulators on apricot pollen germination and pollen tube growth. [Method] Pollens of six kinds of Xinjiang apricots were cultured in solid media supplemented with five plant growth regulators (GA3 , NAA, 2, 4-D, 6-BA, IAA). Then the rate of pollen germination and the length of pollen tube were respectively measured. [Result] In a certain concentration range, GA3 most significantly promoted the pollen germination and the pollen tube growth of Shushanggan, Kalayulvke, Dayoujia, Yiliakeyulvke and Kabakehuanna; NNA had the strongest improvement function on Kumaiti’s pollen germination and pollen tube growth. [Conclusion] All the five plant growth regulators promoted the pollen germination and the pollen tube growth of apricots at low concentration but inhibited them at high concentration.展开更多
Endo-β-glucanases play vital roles in the regulation of pollen tube growth. Here, a previously identified endo-1,4-β-glucanase from Lilium longiflorum (lily), named LlpCel1, was expressed in Escherichia coli, purifi...Endo-β-glucanases play vital roles in the regulation of pollen tube growth. Here, a previously identified endo-1,4-β-glucanase from Lilium longiflorum (lily), named LlpCel1, was expressed in Escherichia coli, purified, and further investigated for its physiological function. The recombinant LlpCel1 protein hydrolyzed carboxy-methylcellulose (CMC) and exhibited activity towards laminarin from Eisenia. arborea and 1,3:1,4-β-glucan of barley. The pH for the optimum activity was 6.0 and the value of Km calculated from CMC was 5.0 mg/mL. Adding EDTA resulted in the total loss of the enzymatic activity, and this effect could be restored by the addition of Ca2+. Western blotting analysis showed that LlpCel1 protein was present in pollen grains and rehydrated pollen grains, and the amount of the protein was increased during pollen germinating, but not in the pollen tube. Consistently, the immunofluorescence labeling study with the antibody against LlpCel1 also indicated the presence of LlpCel1 at the begin-ning of germination, but not in the elongating pollen tube. Furthermore, incubation of LlpCel1 with pollen at the beginning of pollen germination increased the germination percentage and the length of pollen tube. All of these results suggested that LlpCel1 could play an important role in the regulation of lily pollen germination and the initiation of pollen tube growth.展开更多
Double fertilization in angiosperms requires the targeted delivery of immotile sperm to the eggs through pollen tubes. The polarity of tip-growing pollen tubes is maintained through dynamic association of active Rho G...Double fertilization in angiosperms requires the targeted delivery of immotile sperm to the eggs through pollen tubes. The polarity of tip-growing pollen tubes is maintained through dynamic association of active Rho GTPases of plants (ROP-GTP) with the apical plasma membrane. Guanine nucleotide exchange factors for ROPs (RopGEFs) catalyze the activation of ROPs and thereby affect spatiotemporal ROP signaling. Whereas RopGEFs have been found to be phosphorylated proteins, the kinases responsible for their phos- phorylation in v/vo and biological consequences of RopGEF phosphorylation in pollen tube growth remain unclear. We report here that the Arabidopsis AGC1.5 subfamily of cytoplasmic kinases is critical for the restricted localization of ROP-GTP during pollen tube growth. Loss of AGCI.5 and AGC1.7 functions resulted in the mistargeting of active ROPs and defective events downstream of ROP signaling in pollen tubes. AGCI.5 interacts with RopGEFs via their catalytic PRONE domain and phosphorylates RopGEFs at a conserved Ser residue of PRONE domain. Loss of AGC1.5 and AGC1.7 functions resulted in the mistargeting of RopGEFs in pollen tubes, similar to the phenotype caused by the mutation that renders RopGEFs non-phosphorylatable by AGC1.5. Collectively, our results provide mechanistic insights into the spatiotem- poral activation of ROPs during the polar growth of pollen tubes.展开更多
In this article, the functions of D-myo-inositol-1,4,5-trisphosphate (Ins [1,4,5] P3) in regulating pollen tube polar growth were investigated by application of caged version of the phosphoinositides. To increase the ...In this article, the functions of D-myo-inositol-1,4,5-trisphosphate (Ins [1,4,5] P3) in regulating pollen tube polar growth were investigated by application of caged version of the phosphoinositides. To increase the intracellular Ins(1,4,5)P3 concentration at the apical region of pollen tube, the caged Ins(1,4,5)P3 loaded by osmotic shock was activated by 10 s 360 nm UV flash at this domain (10 μm from the tip). More than 70% pollen tubes were induced swelling at apex and/or growth axis reorientation, accompanying temporarily growth arrest, by localized increase of Ins(1,4,5)P3 concentration (n=21). While, pollen tubes without being loaded with caged Ins(1,4,5)P3 had not response to the same dosage UV flash. With FM 1-43 fluorescent staining, it was found that growth perturbation by UV activated caged Ins(1,4,5)P3 had tight link with membrane trafficking at the apical zone of pollen tubes. Upon UV pulse, the apical V-shaped bright area where was full of secretory vesicles spread to a much broader region, which implied that actin filaments at the apical region were remodeled. It was also observed that the FM 1-43 fluorescence intensity at tip remarkably increased than that before UV flash, which demonstrated that more secretory vesicles were accumulated at this region. To estimate the role of Ins(1,4,5)P3 in modulating intracellular calcium concentration and distribution, dextran conjugated fluorescent dye Calcium Green-1 and Rhodamine B were microinjected into pollen tubes together with caged Ins(1,4,5)P3. The results showed that calcium concentration at the subapical region increased upon UV released Ins(1,4,5)P3. Consequently, the tip-focused calcium gradient under the apical dome of pollen tube was disturbed. Simultaneously, the pollen tube bulged at the apical region and its growth rate decreased. As the tip-focused calcium gradient was reestablished, the pollen tube morphology and growth recovered to the normal level. Therefore, Ins(1,4,5)P3 can modulate pollen tube growth rate and direction through mobilizing intracellular calcium and regulating vesicle trafficking during pollen tube finding its way to the ovary.展开更多
We examined the influence of storage time on germinability and tube growth of freeze stored pollens collected from 25 wild male plants in Actinidia eriantha variety. Pollens were stored in freezer at - 20°C ...We examined the influence of storage time on germinability and tube growth of freeze stored pollens collected from 25 wild male plants in Actinidia eriantha variety. Pollens were stored in freezer at - 20°C for six months and one year periods to determine changing at germinability in time. In vitro germination was conducted in certain cultural medium defined for Actinidia genus. The results showed that the germination percentages and tube lengths of genotypes decreased at the end of storage period. MH22, MH45, MH47, MH56, MH67, MH70, MH71, MH72, MH74, MH55 and MH61 genotypes were evaluated as vigor genotypes, because they maintained their viability and germination capability displaying statistically insignificant decreasing although their tube lengths significantly decreased except MH67. This investigation provided to determine some robust wild male germplasm resources in A. eriantha in point of durability of pollens against long term conservation for using at future pollination and breeding programs.展开更多
In angiosperms,pollen tube growth is critical for double fertilization and seed formation.Many of the factors involved in pollen tube tip growth are unknown.Here,we report the roles of pollenspecific GLYCEROPHOSPHODIE...In angiosperms,pollen tube growth is critical for double fertilization and seed formation.Many of the factors involved in pollen tube tip growth are unknown.Here,we report the roles of pollenspecific GLYCEROPHOSPHODIESTER PHOSPHO DIESTERASE-LIKE(GDPD-LIKE)genes in pollen tube tip growth.Arabidopsis thaliana GDPD-LIKE6(At GDPDL6)and At GDPDL7 were specifically expressed in mature pollen grains and pollen tubes and green fluorescent protein(GFP)-At GDPDL6 and GFP-At GDPDL7 fusion proteins were enriched at the plasma membrane at the apex of forming pollen tubes.Atgdpdl6 Atgdpdl7 double mutants displayed severe sterility that was rescued by genetic complementation with At GDPDL6 or At GDPDL7.This sterility was associated with defective male gametophytic transmission.Atgdpdl6 Atgdpdl7 pollen tubes burst immediately after initiation of pollen germination in vitro and in vivo,consistent with the thin and fragile walls in their tips.Cellulose deposition was greatly reduced along the mutant pollen tube tip walls,and the localization of pollen-specific CELLULOSE SYNTHASE-LIKE D1(CSLD1)and CSLD4 was impaired to the apex of mutant pollen tubes.A rice pollen-specific GDPD-LIKE protein also contributed to pollen tube tip growth,suggesting that members of this family have conserved functions in angiosperms.Thus,pollen-specific GDPDLIKEs mediate pollen tube tip growth,possibly by modulating cellulose deposition in pollen tube walls.展开更多
基金the National Natural Science Foundation of China (Grants Nos. 30070454 and 30471045)
文摘The germination of pollen grain in vitro and the growth of pollen tube in the pistil of rice were observed with a microscope. The stigma was removed at different time points after pollination to study its effect on seed setting rate. The rice pollen grain started to germinate at 2 min after pollination and the pollen tube penetrated stigma into style in 5-10 min, 30 min later the end of pollen tube reached the bottom of ovary, and only some pollen tubes arrived at embryo sac at 40 min after pollination. Meanwhile, a small amount of callose began to deposit in the pollen tubes, a great deal of callose was observed at 50 min after pollination, whereas the pollen grain began to shrink. The growing rates of pollen tube in the rice stigma, style and ovary were 1500, 5000, and 5400 μm/h, respectively. The seed setting rate was quite low when the stigma was removed at about 10-15 min after pollination, gradually increased when it removed at 20 min to 50 min after pollination, and over 60% when it removed at 50 min after pollination and finally tended to be stable.
基金supported by grants from the National Research Foundation, Ministry of Education, Science and Technology (2019R1C1C1002636 and RS-2023-00217064 to Y.-J.K., 2021R1A4A2001968 to S.T.K., and 2021M3E5E6025387 and 2021R1A5A1032428 to K.-H.J.)the New Breeding Technologies Development Program (PJ01661002 to K.-H.J.)the Rural Development Administration, Republic of Korea。
文摘Pollen tube growth is essential for successful double fertilization, which is critical for grain yield in crop plants. Rapid alkalinization factors(RALFs) function as ligands for signal transduction during fertilization. However, functional studies on RALF in monocot plants are lacking.Herein, we functionally characterized two pollen-specific RALFs in rice(Oryza sativa) using multiple clustered regularly interspaced palindromic repeats(CRISPR)/CRISPR-associated protein 9-induced loss-of-function mutants,peptide treatment, expression analyses, and tag reporter lines. Among the 41 RALF members in rice, OsRALF17 was specifically expressed at the highest level in pollen and pollen tubes.Exogenously applied OsRALF17 or OsRALF19peptide inhibited pollen tube germination and elongation at high concentrations but enhanced tube elongation at low concentrations, indicating growth regulation. Double mutants of OsRALF17 and OsRALF19(ralf17/19) exhibited almost full male sterility with defects in pollen hydration, germination, and tube elongation,which was partially recovered by exogenous treatment with OsRALF17 peptide. This study revealed that two partially functionally redundant OsRALF17 and OsRALF19 bind to Oryza sativa male-gene transfer defective 2(OsMTD2) and transmit reactive oxygen species signals for pollen tube germination and integrity maintenance in rice. Transcriptomic analysis confirmed their common downstream genes, in osmtd2 and ralf17/19. This study provides new insights into the role of RALF, expanding our knowledge of the biological role of RALF in regulating rice fertilization.
基金Open access funding provided by Shanghai Jiao Tong Universitysupported by the Natural Science Foundation of China(32172511)+2 种基金the National Key Research and Development Program of China(2022YFD1200501)the Nanjing Agricultural Science and Technology Industry-University-Research Collaboration Project(2022RHCXY)the Earmarked Fund for the China Agriculture Research System(CARS-28).
文摘Due to self-incompatibility(SI)prevents self-fertilization,natural or artificial cross-pollination has been conducted in many orchards to stabilize fruit yield.However,it is still puzzled which routes of self S-RNase arresting pollen tube growth.Herein,17 COBRA genes were isolated from pear genome.Of these genes,the pollen-specifically expressed PbCOB.A.1 and PbCOB.A.2 positively mediates pollen tube growth.The promoters of PbCOB.A.1 and/or PbCOB.A.2 were bound and activated by PbABF.E.2(an ABRE-binding factor)and PbC2H2.K16.2(a C2H2-type zinc finger protein).Notably,the expressions of PbCOB.A.1,PbCOB.A.2,and PbC2H2.K16.2 were repressed by self S-RNase,suggesting that self S-RNase reduces the expression of PbCOB.A.1 and PbCOB.A.2 by decreasing the expression of their upstream factors,such as PbC2H2.K16.2,to arrest pollen tube growth.PbCOB.A.1 or PbCOB.A.2 accelerates the growth of pollen tubes treated by self S-RNase,but can hardly affect level of reactive oxygen species and deploymerization of actin cytoskeleton in pollen tubes and cannot physically interact with any reported proteins involved in SI.These results indicate that PbCOB.A.1 and PbCOB.A.2 may not relieve S-RNase toxicity in incompatible pollen tube.The information provides a new route to elucidate the arresting pollen tube growth during SI reaction.
基金Supported by the State Key Basic Research and Development Plan of China(2004CB117306)
文摘With radical global climate change and global warming, high temperature stress has become one of major factors exerting a major Influence on crop production, In the cotton (Gossyplum hirsutum L.)-growlng areas of China, especially in the Yangtze River valley, unexpected periodic episodes of extreme heat stress usually occur In July and August, the peak time of cotton flowering and boll loading, resulting In lower boll set and lint yield. Breeding programs for screening high temperature-tolerant cotton germplasm and cultlvars are urgent In order to stabilize yield in the current and future warmer weather conditions. In the present study, 14 cotton cultivars were quantified for in vitro pollen germination and pollen tube growth in response to temperatures ranging from 10 to 50 ℃ at 5 ℃ intervals. Different cotton genotypes varied In their in vitro pollen germination and pollen tube length responses to the different temperatures. Maximum pollen germination and pollen tube length ranged from 25.2% to 56.2% and from 414 to 682 pro, respectively. The average cardinal temperatures (Tmin,, Topt, and Tmax) also varied among the 14 cultivars and were 11.8, 27.3, and 42.7 ℃ for pollen germination and 11.8, 27.8, and 44.1 ℃ for maximum pollen tube length. Variations In boll retention and boll numbers per plant in field experiments were found for the 14 cotton cultivars and the boll retention and boll retained per plant on 20 August varied considerably In different years according to weather conditions. Boll retention on 20 August was highly correlated with maximum pollen germination (R^2 = 0.84) and pollen tube length (R^2=0.64). A screening method based on principle component analysis of the combination of pollen characterlatics In an in vitro experiment and boll retention testing In the field environment was used In the present study and, as a result, the 14 cotton cultlvars could be classified as tolerant, moderately tolerant, moderately susceptible and susceptible to high temperature.
文摘Ca2+ has been established as an important second messenger regulating pollen germination and tube growth. However, to date, only a few signaling components have been identified to decode and relay Ca2+ signals in growing pollen tubes. Here, we report a function for the calcineurin B-like (CBL) Ca2+ sensor proteins CBL1 and CBL9 from Arabidopsis in pollen germination and tube growth. Both proteins are expressed in mature pollen and pollen tubes and impair pollen tube growth and morphology if transiently overexpressed in tobacco pollen. The induction of these phenotypes requires efficient plasma membrane targeting of CBL1 and is independent of Ca2+ binding to the fourth EF-hand of CBL1. Overexpression of CBL1 or its closest homolog CBL9 in Arabidopsis renders pollen germination and tube growth hypersensitive towards high external K+ concentrations while disruption of CBL1 and CBL9 reduces pollen tube growth under low K~ conditions. Together, our data identify a crucial function for CBL1 and CBL9 in pollen germination and tube growth and suggest a model in which both proteins act at the plasma membrane through regulation of K+ homeostasis.
基金This work was supported by the National Basic Research Program (2014CB954203), the National Natural Science Foundation of China (31670180, 31470283), and the Fundamental Research Funds for the Cen- tral Universities (Izujbky-2016-bt05) to Y.X.
文摘Dynamics of the actin cytoskeleton are essential for pollen germination and pollen tube growth. ACTIN- DEPOLYMERIZlNG FACTORs (ADFs) typically contribute to actin turnover by severing/depolymerizing actin filaments. Recently, we demonstrated that Arabidopsis subclass III ADFs (ADF5 and ADF9) evolved F-actin-bundling function from conserved F-actin-depolymerizing function. However, little is known about the physiological function, the evolutional significance, and the actin-bundling mechanism of these neo- functionalized ADFs. Here, we report that loss of ADF5 function caused delayed pollen germination, retarded pollen tube growth, and increased sensitive to latrunculin B (LatB) treatment by affecting the generation and maintenance of actin bundles. Examination of actin filament dynamics in living cells revealed that the bundling frequency was significantly decreased in adf5 pollen tubes, consistent with its biochem- ical functions. Further biochemical and genetic complementation analyses demonstrated that both the N- and C-terminal actin-binding domains of ADF5 are required for its physiological and biochemical functions. Interestingly, while both are atypical actin-bundling ADFs, ADF5, but not ADF9, plays an important role in mature pollen physiological activities. Taken together, our results suggest that ADF5 has evolved the function of bundling actin filaments and plays an important role in the formation, organization, and maintenance of actin bundles during pollen germination and pollen tube growth.
基金Supported by the National Natural Science Foundation of China (No. 30170090) and partially financed as a Joint Research Program by the Japanese Society for the Promotion of Science and the National Natural Science Foundation of China
文摘The present study is expected to provide better understanding of how exo-β-glucanase acts on the biochemo-rheological property of the cell wall, and thus the regulation mechanism of pollen tube growth. A polymerase chain reaction (PCR)-based strategy and cDNA library screening were used to clone an exo-β-glucanase gene (LP-Exol) from Lilium Iongiflorum pollen tubes. The LP-Exol sequence showed a high level of homology to that of reported plant exo-β-glucan hydrolases. Phylogenetic analysis demonstrated that there was close relationship between LP-Exol and other exo-glucan hydrolases in Poacea plants. Northern analysis indicated that LP-Exol transcripts stored in pollen grains at low levels; LP-Exol transcription level enhanced at 2 h after the onset of pollen incubation and remained relatively high during pollen tube elongation. Moreover, the expression of LP-Exol was undetectable in petals, filaments, stigmas, styles, ovaries, leaves, and stems. These results suggest that LP-Exol gene is a late pollen gene, specifically contributing to the regulation of lily pollen germination and pollen tube growth.
基金Supported by the FIRST Research Program of the University of Milan
文摘Although pollen tube growth is a prerequisite for higher plant fertilization and seed production, the processes leading to pollen tube emission and elongation are crucial for understanding the basic mechanisms of tip growth. It was generally accepted that pollen tube elongation occurs by accumulation and fusion of Golgi-derived secretory vesicles (SVs) in the apical region, or clear zone, where they were thought to fuse with a restricted area of the apical plasma membrane (PM), defining the apical growth domain. Fusion of SVs at the tip reverses outside cell wall material and provides new segments of PM. However, electron microscopy studies have clearly shown that the PM incorporated at the tip greatly exceeds elongation and a mechanism of PM retrieval was already postulated in the mid-nineteenth century. Recent studies on endocytosis during pollen tube growth showed that different endocytic pathways occurred in distinct zones of the tube, including the apex, and led to a new hypothesis to explain vesicle accumulation at the tip; namely, that endocytic vesicles contribute substantially to V-shaped vesicle accumulation in addition to SVs and that exocytosis does not involve the entire apical domain. New insights suggested the intriguing hypothesis that modulation between exo- and endocytosis in the apex contributes to maintain PM polarity in terms of lipid/protein composition and showed distinct degradation pathways that could have different functions in the physiology of the cell. Pollen tube growth in vivo is closely regulated by interaction with style molecules. The study of endocytosis and membrane recycling in pollen tubes opens new perspectives to studying pollen tube-style interactions in vivo.
基金Supported by Key Technology Integration and Demonstration of Xinjiang Characteristic Fruit Trees'High Efficiency and Safe Production,Science and Technical Plan Project of Xinjiang Uygur Autonomous Region(201130102)Key Technology Integration and Demonstration of Xinjiang Apricot Industrial Development,Science and Technical Plan Project of Xinjiang Uygur Autonomous Region(200931101)Financial Support from Xinjiang Uygur Autonomous Region Fruit Trees Key Subject~~
文摘[Objective] This study aimed to investigate the influence of different plant growth regulators on apricot pollen germination and pollen tube growth. [Method] Pollens of six kinds of Xinjiang apricots were cultured in solid media supplemented with five plant growth regulators (GA3 , NAA, 2, 4-D, 6-BA, IAA). Then the rate of pollen germination and the length of pollen tube were respectively measured. [Result] In a certain concentration range, GA3 most significantly promoted the pollen germination and the pollen tube growth of Shushanggan, Kalayulvke, Dayoujia, Yiliakeyulvke and Kabakehuanna; NNA had the strongest improvement function on Kumaiti’s pollen germination and pollen tube growth. [Conclusion] All the five plant growth regulators promoted the pollen germination and the pollen tube growth of apricots at low concentration but inhibited them at high concentration.
基金Supported by the National Natural Science Foundation of China (Grant No. 30170090)
文摘Endo-β-glucanases play vital roles in the regulation of pollen tube growth. Here, a previously identified endo-1,4-β-glucanase from Lilium longiflorum (lily), named LlpCel1, was expressed in Escherichia coli, purified, and further investigated for its physiological function. The recombinant LlpCel1 protein hydrolyzed carboxy-methylcellulose (CMC) and exhibited activity towards laminarin from Eisenia. arborea and 1,3:1,4-β-glucan of barley. The pH for the optimum activity was 6.0 and the value of Km calculated from CMC was 5.0 mg/mL. Adding EDTA resulted in the total loss of the enzymatic activity, and this effect could be restored by the addition of Ca2+. Western blotting analysis showed that LlpCel1 protein was present in pollen grains and rehydrated pollen grains, and the amount of the protein was increased during pollen germinating, but not in the pollen tube. Consistently, the immunofluorescence labeling study with the antibody against LlpCel1 also indicated the presence of LlpCel1 at the begin-ning of germination, but not in the elongating pollen tube. Furthermore, incubation of LlpCel1 with pollen at the beginning of pollen germination increased the germination percentage and the length of pollen tube. All of these results suggested that LlpCel1 could play an important role in the regulation of lily pollen germination and the initiation of pollen tube growth.
基金This research was supported by the National Natural Science Foundation of China (31471304 to Y.Z. 31771558 to S.L.), Natural Science Foundation of Shandong Province (ZR2014CM027 to S.L.), China Postdoctoral Science Foundation (2018M632698 to E.L.), and Research Grants Council of Hong Kong (AoE/M-05/12 and C4011-14R). Y.Z. was partially supported by the Tai-Shan Scholar program from the Shandong Provinciak Government.
文摘Double fertilization in angiosperms requires the targeted delivery of immotile sperm to the eggs through pollen tubes. The polarity of tip-growing pollen tubes is maintained through dynamic association of active Rho GTPases of plants (ROP-GTP) with the apical plasma membrane. Guanine nucleotide exchange factors for ROPs (RopGEFs) catalyze the activation of ROPs and thereby affect spatiotemporal ROP signaling. Whereas RopGEFs have been found to be phosphorylated proteins, the kinases responsible for their phos- phorylation in v/vo and biological consequences of RopGEF phosphorylation in pollen tube growth remain unclear. We report here that the Arabidopsis AGC1.5 subfamily of cytoplasmic kinases is critical for the restricted localization of ROP-GTP during pollen tube growth. Loss of AGCI.5 and AGC1.7 functions resulted in the mistargeting of active ROPs and defective events downstream of ROP signaling in pollen tubes. AGCI.5 interacts with RopGEFs via their catalytic PRONE domain and phosphorylates RopGEFs at a conserved Ser residue of PRONE domain. Loss of AGC1.5 and AGC1.7 functions resulted in the mistargeting of RopGEFs in pollen tubes, similar to the phenotype caused by the mutation that renders RopGEFs non-phosphorylatable by AGC1.5. Collectively, our results provide mechanistic insights into the spatiotem- poral activation of ROPs during the polar growth of pollen tubes.
文摘In this article, the functions of D-myo-inositol-1,4,5-trisphosphate (Ins [1,4,5] P3) in regulating pollen tube polar growth were investigated by application of caged version of the phosphoinositides. To increase the intracellular Ins(1,4,5)P3 concentration at the apical region of pollen tube, the caged Ins(1,4,5)P3 loaded by osmotic shock was activated by 10 s 360 nm UV flash at this domain (10 μm from the tip). More than 70% pollen tubes were induced swelling at apex and/or growth axis reorientation, accompanying temporarily growth arrest, by localized increase of Ins(1,4,5)P3 concentration (n=21). While, pollen tubes without being loaded with caged Ins(1,4,5)P3 had not response to the same dosage UV flash. With FM 1-43 fluorescent staining, it was found that growth perturbation by UV activated caged Ins(1,4,5)P3 had tight link with membrane trafficking at the apical zone of pollen tubes. Upon UV pulse, the apical V-shaped bright area where was full of secretory vesicles spread to a much broader region, which implied that actin filaments at the apical region were remodeled. It was also observed that the FM 1-43 fluorescence intensity at tip remarkably increased than that before UV flash, which demonstrated that more secretory vesicles were accumulated at this region. To estimate the role of Ins(1,4,5)P3 in modulating intracellular calcium concentration and distribution, dextran conjugated fluorescent dye Calcium Green-1 and Rhodamine B were microinjected into pollen tubes together with caged Ins(1,4,5)P3. The results showed that calcium concentration at the subapical region increased upon UV released Ins(1,4,5)P3. Consequently, the tip-focused calcium gradient under the apical dome of pollen tube was disturbed. Simultaneously, the pollen tube bulged at the apical region and its growth rate decreased. As the tip-focused calcium gradient was reestablished, the pollen tube morphology and growth recovered to the normal level. Therefore, Ins(1,4,5)P3 can modulate pollen tube growth rate and direction through mobilizing intracellular calcium and regulating vesicle trafficking during pollen tube finding its way to the ovary.
文摘We examined the influence of storage time on germinability and tube growth of freeze stored pollens collected from 25 wild male plants in Actinidia eriantha variety. Pollens were stored in freezer at - 20°C for six months and one year periods to determine changing at germinability in time. In vitro germination was conducted in certain cultural medium defined for Actinidia genus. The results showed that the germination percentages and tube lengths of genotypes decreased at the end of storage period. MH22, MH45, MH47, MH56, MH67, MH70, MH71, MH72, MH74, MH55 and MH61 genotypes were evaluated as vigor genotypes, because they maintained their viability and germination capability displaying statistically insignificant decreasing although their tube lengths significantly decreased except MH67. This investigation provided to determine some robust wild male germplasm resources in A. eriantha in point of durability of pollens against long term conservation for using at future pollination and breeding programs.
基金supported by the Fundamental Research Funds for the Central Universities(2572021DT01)the National Natural Science Foundation of China(31770637)+1 种基金the Innovation Project of State Key Laboratory of Tree Genetics and Breeding(2014A01)the Heilongjiang Touyan Innovation Team Program(Tree Genetics and Breeding Innovation Team)。
文摘In angiosperms,pollen tube growth is critical for double fertilization and seed formation.Many of the factors involved in pollen tube tip growth are unknown.Here,we report the roles of pollenspecific GLYCEROPHOSPHODIESTER PHOSPHO DIESTERASE-LIKE(GDPD-LIKE)genes in pollen tube tip growth.Arabidopsis thaliana GDPD-LIKE6(At GDPDL6)and At GDPDL7 were specifically expressed in mature pollen grains and pollen tubes and green fluorescent protein(GFP)-At GDPDL6 and GFP-At GDPDL7 fusion proteins were enriched at the plasma membrane at the apex of forming pollen tubes.Atgdpdl6 Atgdpdl7 double mutants displayed severe sterility that was rescued by genetic complementation with At GDPDL6 or At GDPDL7.This sterility was associated with defective male gametophytic transmission.Atgdpdl6 Atgdpdl7 pollen tubes burst immediately after initiation of pollen germination in vitro and in vivo,consistent with the thin and fragile walls in their tips.Cellulose deposition was greatly reduced along the mutant pollen tube tip walls,and the localization of pollen-specific CELLULOSE SYNTHASE-LIKE D1(CSLD1)and CSLD4 was impaired to the apex of mutant pollen tubes.A rice pollen-specific GDPD-LIKE protein also contributed to pollen tube tip growth,suggesting that members of this family have conserved functions in angiosperms.Thus,pollen-specific GDPDLIKEs mediate pollen tube tip growth,possibly by modulating cellulose deposition in pollen tube walls.