Characterization,expression dynamics,and potential function of OPA1 for regulation of mitochondrial morphology during spermiogenesis in Phascolosoma esculenta

Mitochondria undergo morphological changes during spermatogenesis in some animals.The mechanism and role of mitochondrial morphology regulation,however,remain somewhat unclear.In this study,we analyzed the molecular characteristics,expression dynamics and subcellular localization of optic atrophy protein 1(OPA1),a mitochondrial fusion and cristae maintenance-related protein,to reveal the possible regulatory mechanisms underlying mitochondrial morphology in Phascolosoma esculenta spermiogenesis.The full-length cDNA of the P.esculenta opa1 gene(Pe-opa1)is 3743 bp in length and encodes 975 amino acids.The Pe-OPA1 protein is highly conservative and includes a transmembrane domain,a GTPase domain,two helical bundle domains,and a lipid-interacting stalk.Gene and protein expression was higher in the coelomic fluid(a site of spermatid development)of male P.esculenta and increased first and then decreased from March to December.Moreover,their expression during the breeding stage was significantly higher than during the non-breeding stage,suggesting that Pe-OPA1 is involved in P.esculenta reproduction.The Pe-OPA1 protein was more abundant in components consisting of many spermatids than in components without,indicating that Pe-OPA1 mainly plays a role in the spermatid in coelomic fluid.Moreover,Pe-OPA1 was mainly detected in the spermatid mitochondria.Immunofluorescence experiments showed that the Pe-OPA1 are constitutively expressed and co-localized with mitochondria during spermiogenesis,suggesting its involvement in P.esculenta spermiogenesis.These results provide evidence for Pe-OPA1's involvement in the regulation of mitochondrial morphology during spermiogenesis.

Germ cell-specific deletion of Pex3 reveals essential roles of PEX3-dependent peroxisomes in spermiogenesis

Peroxisomes are organelles enclosed by a single membrane and are present in various species.The abruption of peroxisomes is correlated with peroxisome biogenesis disorders and single peroxisomal enzyme deficiencies that induce diverse diseases in different organs.However,little is known about the protein compositions and corresponding roles of heterogeneous peroxisomes in various organs.Through transcriptomic and proteomic analyses,we observed heterogenous peroxisomal components among different organs,as well as between testicular somatic cells and different developmental stages of germ cells.As Pex3 is expressed in both germ cells and Sertoli cells,we generated Pex3 germ cell-and Sertoli cell-specific knockout mice.While Pex3 deletion in Sertoli cells did not affect spermatogenesis,the deletion in germ cells resulted in male sterility,manifested as the destruction of intercellular bridges between spermatids and the formation of multinucleated giant cells.Proteomic analysis of the Pex3-deleted spermatids revealed defective expressions of peroxisomal proteins and spermiogenesis-related proteins.These findings provide new insights that PEX3-dependent peroxisomes are essential for germ cells undergoing spermiogenesis,but not for Sertoli cells.

The Study on the Localization of Acid Phosphatase Activity During Spermiogenesis in Chinese Mitten-Handed Crab,Eriocheir sinensis

Ultrastructural cytochemical techniques and electron microscopy were used for localization of acid phosphatase activity during spermiogenesis in Eriocheir sinemsis. The results showed that： Acid phosphatase was synthesized in the endoplasmic reticulum in the early spermatids. The acid phosphatase was found gradually in nucleus, the membrane of acrosomal vesicle, the cytoplasmic region and the acrosomal tubule. And then the reaction product particles became thicker during the spermiogenesis. In the mature sperm, acid phosphatase was localized in the percutor organ slightly, but it was massive and compact in the acrosomal tubule.

The cytoplasmic droplet may be indicative of sperm motility and normal spermiogenesis

Although the cytoplasm of spermatids is removed at the end of spermiogenesis, a tiny portion is usually retained in the sperm flagellum, which is termed the cytoplasmic droplet （CD） in mammals. CDs are believed to play a role in sperm volume adaptation. However, we have noticed that epididymal spermatozoa that display initial （flagellation in situ） and progressive motility mostly possess CDs, whereas spermatozoa without CDs are rarely motile, suggesting that CDs have a role in motility development during sperm epididymal maturation. In the present study, we analyzed the relationship between the presence or absence of CDs, motility development and positional changes of CDs during sperm epididymal maturation in mice and monkeys. We also examined CDs on spermatozoa of three knockout mouse lines with late spermiogenic defects. Our data suggest that the CD is a normal organelle transiently present exclusively on epididymal spermatozoa, and normal CD morphology and location are associated with normal motility development during epididymal maturation of spermatozoa. Abnormal CD formation, e.g., a complete lack of CDs or ectopic CDs, is indicative of defective spermiogenesis. If CDs are essential for sperm motility development, then CDs may represent an ideal drug target for the development of non-hormonal male contraceptives.

Intramanchette transport during primate spermiogenesis： expression of dynein, myosin Va, motor recruiter myosin Va, VIIa-Rab27a/b interacting protein, and Rab27b in the manchette during human and monkey spermiogenesis

Aim： To show whether molecular motor dynein on a microtubule track, molecular motor myosin Va, motor recruiter myosin Va, VIIa-Rab27a/b interacting protein （MyRIP）, and vesicle receptor Rab27b on an F-actin track were present during human and monkey spermiogenesis involving intramanchette transport （IMT）. Methods： Spermiogenic cells were obtained from three men with obstructive azoospermia and normal adult cynomolgus monkey （Macacafascieularis）. Immunocytochemical detection and reverse transcription-polymerase chain reaction （RT-PCR） analysis of the proteins were carried out. Samples were analyzed by light microscope. Results： Using RT-PCR, we found that dynein, myosin Va, MyRIP and Rab27b were expressed in monkey testis. These proteins were localized to the manchette, as shown by immunofluorescence, particularly during human and monkey spermiogenesis. Conclusion： We speculate that during primate spermiogenesis, those proteins that compose microtubule-based and actin-based vesicle transport systems are actually present in the manchette and might possibly be involved in intramanchette transport.

A Novel Type of Spermiogenesis in the Nile Catfish Chrysichthys auratus (Siluriformes: Bagridae) in Egypt, with Description of Spermatozoon Ultrastructure

Spermiogenesis and spermatozoon ultrastructure in the Nile catfish Chrysichthys auratus are described using transmission electron microscopy. Spermiogenesis involves some unique peculiarities such as ： the development of the centriolar complex and the initial segment of the flagellum in a position directly perpendicular to the basal pole of the nucleus, as a result of absence of nuclear rotation ; lack of a cytoplasmic canal during differentiation of the spermatids into spermatozoa; the base of the basal body is not traversed by the basal plate; a basal foot anchors the basal body to the nucleus; and the presence of numerous vesicles around the midpiece and base of the flagellum. In addition, spermiogenesis includes some common features such as： chromatin compaction; formation of a medial shallow nuclear fossa; and elimination of excess cytoplasm. The mature spermatozoon has an elongate conical-shaped head with no acrosome or acrosomal vesicle, a long midpiece with numerous vesicles that continue backwards around the base of the flagellum and a long tail or flagellum, which has no lateral fins or a membranous compartment. The mitochondria lie close to the nucleus basal pole and surround the initial segment of the axoneme and are separated from the flagellum by the inner mitochondrial envelope due to disappearance of the cytoplasmic canal. The flagellum has the classical axoneme structure of a 9 ＋ 2 microtubular pattern. On the basis of the peculiar features mentioned above, it is concluded that spermiogenesis in this Nile catfish is a synapomorphic type derived from types Ⅰ and Ⅱ spermiogenesis, which are common among teleosts. Accordingly, this type could be considered as a novel type of spermiogenesis and could be termed as ＂type Ⅲ＂.

Molecular Characterization, Tissue Distribution and Localization of Larimichthys crocea Kif3a and Kif3b and Expression Analysis of Their Genes During Spermiogenesis

KIF3A and KIF3B are two N-terminal motor proteins belonging to the kinesin-II superfamily that play essential roles in spermiogenesis.To understand the roles played by KIF3 A/3B during spermatogenesis of large yellow croaker Larimichthys crocea,we studied the testis characteristics at different developmental stages of L.crocea,and determined the spatiotemporal expression patterns of kif3a and kif3b during spermiogenesis.Quantitative real-time PCR(qR T-PCR)showed that the overall trends of kif3 a/3 b m RNA abundance during testis development are similar.From stage Ⅱ to stage V,kif3a/3b m RNA abundances first increased and then fell after reaching a peak at stage IV.Interestingly,the m RNA abundances of both genes at stage V were higher than those at stages Ⅱ and Ⅲ.In addition,it is worth of noting that kif3 b m RNA abundance was higher than that of kif3a at all stages.Fluorescence in situ hybridization results revealed that kif3a/3b m RNA abundance dynamics were consistent with the migration of mitochondria,the deformation of nucleus,and the formation of tail.The m RNA hybridization signals of both genes first appeared either around the nuclear periphery or on the side of the nuclei,then appeared at one side of nuclei,and finally were mainly on the tail during spermiogenesis.Our findings contributed to better understanding the molecular mechanisms of spermiogenesis in fish;and suggested that KIF3A and KIF3B may participate in the intracellular transport of mitochondria,nuclear deformation,and the formation of tail during the spermiogenesis in L.crocea.

miR-199-5p regulates spermiogenesis at the posttranscriptional level via targeting Tekt1 in allotriploid crucian carp

Background:Sperm abnormalities are one of the primary factors leading to male sterility,but their pathogenesis is still unclear.Although miRNAs are suggested to exert important roles in the regulation of spermatogenesis at both transcriptional and posttranscriptional levels,little is currently known regarding the regulation of sperm flagella assembly by microRNAs(miRNAs).The role of miRNAs in the development of sperm abnormalities in sterile triploid fish has not been studied.Results:In this study,we found that miR-199-5p was widely expressed in all detected tissues of different-ploidy crucian carp.As one of the testis-specific candidate markers,Tekt1 was predominantly expressed in the testis.Quantitative real-time PCR(qRT-PCR)analyses showed that the expression trend of miR-199-5p was exactly opposite to that of Tekt1.Through bioinformatics analysis,we identified a putative miR-199-5p binding site in the Tekt1 mRNA.We further identified Tekt1 as a target of miR-199-5p using luciferase reporter assay.Finally,we confirmed that miR-199-5p was necessary for sperm flagellar assembly and spermatogenesis in vivo via intraperitoneal injection of miR-199-5p antagomir or agomir in diploid red crucian carp.Moreover,miR-199-5p gainof-function could lead to spermatids apoptosis and abnormal spermatozoa structure,which is similar to that of allotriploid crucian carp.Conclusions:Our studies suggested that abnormally elevated miR-199-5p inhibited the sperm flagella formation in spermiogenesis by negatively regulating the expression of Tekt1,thereby causing sperm abnormalities of male allotriploid crucian carp.

Evidence for the role of KIF17 in fish spermatid reshaping:expression pattern of KIF17 in Larimichthys polyactis spermiogenesis

The homodimeric kinesin-2 protein KIF17 functions in intracellular transport and spermiogenesis in mammals.However,its role in fish spermiogenesis has not been reported.Here,we aimed to clone full-length kif17 cDNA and determine the molecular characteristics and expression patterns of KIF17 in Larimichthys polyactis spermiogenesis.The full-length cDNA of L.polyactis kif17(Lp-kif17)was sequenced and found to contain a 332-bp 5′untranslated region,480-bp 3′untranslated region,and 2433-bp open reading frame encoding 810 amino acids.Bioinformatics analyses showed that L.polyactis KIF17(Lp-KIF17)shared high sequence similarity with homologs in other animals and possessed an N-terminal motor domain with microtubule-binding sites and adenosine triphosphate(ATP)hydrolysis sites,a stalk domain containing two coiled-coil regions,and a C-terminal tail domain.The Lp-kif17 mRNA was widely expressed in various tissues,with the highest level in the brain,followed by that in the testis.Fluorescence in situ hybridization(FISH)analysis revealed that Lp-kif17 was continuously expressed in spermiogenesis,showing that it had potential functions in this process.Using immunofluorescence(IF)analysis,we found that Lp-KIF17 colocalized with tubulin and was transferred from the perinuclear cytoplasm to the side of spermatid where the tail forms during spermiogenesis.These findings suggested that KIF17 is involved in L.polyactis spermiogenesis.In particular,it may participate in nuclear shaping and tail formation by interacting with perinuclear microtubules during spermatid reshaping.In addition to providing evidence for the role of KIF17 in fish spermatid reshaping,this study provides important data for studies of reproductive biology in L.polyactis.

Mitochondria: transportation, distribution and function during spermiogenesis

Spermiogenesis is a dynamic process which includes organelle reorganization and new structure formation. The morphology and distribution of the mitochondria in germ cells change to accommodate the cellular requirement. Multiple molecular motors and related proteins participate in carrying and anchoring mitochondria to the midpiece during spermiogenesis and this process is regulated precisely. Energetic metabolism provides energy for cellular activity and influences sperm survival and motility directly. Ubiquitination of mitochondria takes place during spermiogenesis, which has been implicated in sperm quality control and mitochondrial inheritance. In light of the essential roles of mitochondria in energy production, calcium homeostasis and apoptosis, mitochondria dysfunction cause severe human diseases, such as male infertility. The present study paves a way for a more detailed exploration of the biology of mitochondria during spermiogenesis.

Molecular Cloning and Putative Functions of KIFC1 for Acrosome Formation and Nuclear Reshaping during Spermiogenesis in Schlegel’s Japanese Gecko Gekko japonicus

Spermiogenesis, occurring in the male testis, is a complicated and highly-ordered developmental process resulting in the production of fertile mature sperm. In Gekko japonicus, this process occurs in 7 steps during which the spermatids undergo dramatic changes in the cytoskeleton and nucleus. Here, we cloned and sequenced the cDNA of the mammalian KIFC1 homologue in the testis of G. japonicus. The 2 344 bp full-length cDNA sequence contained a 191 bp 5’-untranslated region, a 134 bp 3’-untranslated region and a 2 019 bp open reading frame encoding a protein of 672 amino acids. Tissue expression analysis revealed the highest expression of kifc1 mRNA was in the testis. Fluorescence in situ hybridization revealed that the kifc1 mRNA signal was hardly detected in step 1 spermatids but became concentrated at the acrosome of step 2 spermatids and abundant in the nucleus of step 5 spermatids where the nucleus then undergoes dramatic elongation and compression. The kifc1 mRNA signal then gradually disappears in mature sperm. This expression of KIFC1 at specific stages of spermiogenesis in G. japonicus implies its important role in the major cytological transformations such as acrosome biogenesis and nucleus morphogenesis.

Characterization of rtSHSp13 gene encoding a development protein involved in vesicular traffic in spermiogenesis

A cDNA, designated as rtSH3p13, was isolated from a rat testis cDNA library. It consists of 1463 bp nuclear acids,which encodes a protein of 312 amino acids and was assigned the GenBank accession number AF227439. The deduced rtSH3p13 protein is a truncated isoform of SH3p13 as a result of mRNA alternative splicing. It is mainly expressed in the rat testis, detected in spermatids at the steps 8-19 of spermiogenesis, and found around the acrosome. During postnatal development, rtSH3p13 appears on day 18 and reaches maximum on day 60. Further experimental results suggested that rtSH3p13 forms a complex with activated epidermal growth factor receptor (EGFR) and interacts with synaptojanin I. Surprisingly, similar to SH3 domain, the V region of rtSH3p13 also inhibits endocytosis in CHO cells.Our results reveal a link between an rtSH3p13-synaptojanin-clathrin complex-mediated formation of pits and the process of spermiogenesis.

Expression Characteristics and Putative Functions of KIF3A/KIF3B During Spermiogenesis of Phascolosoma esculenta

The microtubule(MT)-associated proteins KIF3A and KIF3B are ubiquitously expressed in a wide range of taxa.This study investigated the functions of these proteins in spermiogenesis,which involves various MT-dependent processes,in Phasco-losoma esculenta.We cloned the complete cDNA of Pe-KIF3A/3B.Structural predictions showed that Pe-KIF3A/3B are composed of a highly conserved motor domain,a coiled-coil domain,and a tail domain.Real-time quantitative PCR showed that Pe-kif3a/3b are expressed in all tissues evaluated,with the highest levels in sperm masses.Fluorescence in situ hybridization and immunofluo-rescence were employed to analyze the dynamic expression patterns of KIF3A/3B during spermiogenesis.Pe-KIF3A/3B consistently co-localized with MTs at all stages of spermiogenesis,indicating their potential functions in cargo trafficking.Pe-KIF3A/3B co-localized with mitochondria,suggesting that they may mediate mitochondrial movement.In late-stage spermatids and mature sperm,co-localization was detected in the midpiece,suggesting that Pe-KIF3A/3B could facilitate midpiece formation.The co-localization of Pe-KIF3A and Pe-KIF3B at all stages of spermiogenesis suggests their function as the heterodimeric KIF3AB.Basing on the ob-served temporal and spatial expression patterns of Pe-KIF3A/3B,MTs,and mitochondria in our study,we suggest that heterodimer KIF3AB has a potential role in acrosome biogenesis,sperm head remodeling,enflagellation,and mitochondrial migration during spermiogenesis in P.esculenta.In addition,our study on the morphological characteristics of spermatogenic cells provided fundamental data on the reproductive biology of P.esculenta.

Ultrastructure Developments During Spermiogenesis in Polydora ciliata(Annelida: Spionidae),a Parasite of Mollusca

Spionid worms of Polydora ciliata inhabit the shells of many commercially important bivalves and cause disease in molluscan aquaculture. Their sperm structure is closely related to their fertilization method. To give an insight into the sperm structure and spermatogenesis, ultrastructure details of the subcellular components of germ cells during spermiogenesis of Polydora ciliata are detected by transmission electron microscopy(TEM). In P. ciliata, during spermiogenesis, chromatin is regularly arranged as dense fibrils and becomes more condensed when the nucleus elongates. Microtubules do not surround the nucleus during its elongation. The Golgi phase is characterized by the formation of proacrosomal granules within the Golgi apparatus. The proacrosomal granules fuse to form a single, spherical acrosomal vesicle that migrates to the anterior pole of the cell. At the time of nuclear condensation, mitochondria become reduced in number but increased in size, causing deep indentation at the base of the nucleus. The mid-piece has a few mitochondria. The cap phase includes the spreading of the acrosomal granule over the surface of the nucleus of the differentiating spermatid. The acrosomal phase of spermiogenesis is typically associated with changes in the shape of the nucleus, acrosome and tail. The relationship of sperm ultrastructure to spermiogenesis in spionidae species was discussed.

SPERMIOGENESIS IN THE YELLOWFIN PORGY (SPARUS LATUS HOUTTUYN), WITH EMPHASIS ON THE ASSOCIATED MITOCHONDRION

Transmission electron microscopy of the yellowfin porgy (Sparus latus Houttuyn) spermatozoa ultrastructure showed the spermatozoon as a primitive type made up of the acrosomeless head , the flagellum , and the midpiece , at the periphery of which was a relatively big mitochondrion with more complex structure . It was found that during spermiogenesis, only one relatively big mitochondrion occurred in both the spermatid and the spermatozoon . This is different from other teleost fishes . During spermiogenesis, the mitochondria number is one, and morphology did not change . All these are different fromthose of other fishes .

CCDC181 is required for sperm flagellum biogenesis and male fertility in mice

The structural integrity of the sperm flagellum is essential for proper sperm function.Flagellar defects can result in male infertility,yet the precise mechanisms underlying this relationship are not fully understood.CCDC181,a coiled-coil domain-containing protein,is known to localize on sperm flagella and at the basal regions of motile cilia.Despite this knowledge,the specific functions of CCDC181 in flagellum biogenesis remain unclear.In this study,Ccdc181 knockout mice were generated.The absence of CCDC181 led to defective sperm head shaping and flagellum formation.Furthermore,the Ccdc181 knockout mice exhibited extremely low sperm counts,grossly aberrant sperm morphologies,markedly diminished sperm motility,and typical multiple morphological abnormalities of the flagella(MMAF).Additionally,an interaction between CCDC181 and the MMAF-related protein LRRC46 was identified,with CCDC181 regulating the localization of LRRC46 within sperm flagella.These findings suggest that CCDC181 plays a crucial role in both manchette formation and sperm flagellum biogenesis.

Cytosolic Ca^(2+)as a multifunctional modulator is required for spermiogenesis in Ascaris suum

The dynamic polar polymers actin fi laments and microtu-bules are usually employed to provide the structural ba-sis for establishing cell polarity in most eukaryotic cells.Radially round and immotile spermatids from nematodes contain almost no actin or tubulin,but still have the abil-ity to break symmetry to extend a pseudopod and initiate the acquisition of motility powered by the dynamics of cytoskeleton composed of major sperm protein(MSP)during spermiogenesis(sperm activation).However,the signal transduction mechanism of nematode sperm activation and motility acquisition remains poorly under-stood.Here we show that Ca^(2+) oscillations induced by the Ca^(2+) release from intracellular Ca^(2+) store through inositol(1,4,5)-trisphosphate receptor are required for Ascaris suum sperm activation.The chelation of cytosolic Ca^(2+) suppresses the generation of a functional pseudopod,and this suppression can be relieved by introducing ex-ogenous Ca^(2+) into sperm cells.Ca^(2+) promotes MSP-based sperm motility by increasing mitochondrial membrane potential and thus the energy supply required for MSP cytoskeleton assembly.On the other hand,Ca^(2+) promotes MSP disassembly by activating Ca^(2+)/calmodulin-depend-ent serine/threonine protein phosphatase calcineurin.In addition,Ca^(2+)/camodulin activity is required for the fusion of sperm-specifi c membranous organelle with the plasma membrane,a regulated exocytosis required for sperm mo-tility.Thus,Ca^(2+)plays multifunctional roles during sperm activation in Ascaris suum.

Rapid Evolution of the Mammalian HILS1 Gene and the Nuclear Condensation Process during Mammalian Spermiogenesis

Many genes associated with reproduction show rapid evolution across diverse animal groups, a result commonly due to adaptive evolution driven by positive selection （Swanson and Vacquier, 2002）. Different theories have been proposed to explain the elevated rates of evolution （Swanson and Vacquier, 2002）, including sperm competition, where sperm compete to fertilize eggs leading to the proteins in the sper- matozoa adaptively evolving to increase their ability to fertilize eggs; sexual conflict, where the egg experiences a loss of fitness when sperm are too abundant; sexual selection, where eggs bind sperm carrying adaptive alleles （Palumbi, 1999）; and cryptic female choice （reviewed in Swanson and Vacquier, 2002）.

Histological and immunocytochemical study of deferens ducts in the Chinese rat snake (Zaocys dhumnades)

To investigate the relationship between structure and function of the deferens ducts in the Chinese rat snake （Zaocys dhumnades）, morphological changes within an annual cycle were observed by routine histological techniques. Also, the correlation of androgen receptor （AR）, estrogen receptor （ER）, progesterone receptor （PR） and aromatase （Ar） expressions in the vas deferens and testis were studied immunohistochemically. To confh＇m that the sperm and the spherical structure existed in deferens ducts, we also used routine histological technique observed deferens ducts in the Striped-tailed rat-snake （Elaphe taeniura）, Red-banded snake （Dinodon rufozonatum）, and Tiger-spotted neck- troughed snake （Rhabdophis tigrina lateralis）. The results showed that the deferens ducts of the Chinese Rat Snake were composed of efferent duct, epididymal duct and vas deferens. Efferent duct contained sperm from August-October, and the sperm were observed in the epididymal duct from August-the following January. Throughout the year （except July） a large number of sperm were present in the vas deferens where a previously unreported spherical structure formed by spermatids was observed, which showed no significant differences in the IOD values of AR-, ER-, PR- and Ar- immunoreactivities. Since the spermatids in the spherical structure were undergoing spermatogenesis and this phenomenon also existed in the Striped-tailed rat-snake and Red-banded snake, the term, seminiferous spherule, was named for this spherical structure This study demonstrated that the testis was the main site for snake spermiogenesis, and the seminiferous spherule in vas deferens was the other Both the epididymis and vas deferens stored sperm; however, the vas deferens was the main organ for sperm storage.

Aquaporins in spermatozoa and testicular germ cells： identification and potential role

Mammalian spermatozoa have relatively high water permeability and swell readily, as in the hypo-osmotic swelling test used in the andrology clinic. Physiologically, spermatozoa experience changes in the osmolality of the surrounding fluids in both the male and the female tracts on their journey from the testis to the ovum. Sperm volume regulation in response to such osmotic challenges is important to maintain a stable cell size for the normal shape and function of the sperm tail. Alongside ion channels for the fluxes of osmolytes, water channels would be crucial for sperm volume regulation. In contrast to the deep knowledge and numerous studies on somatic cell aquaporins （AQPs）, the understanding of sperm AQPs is limited. Among the 13 AQPs, convincing evidence for their presence in spermatozoa has been confined to AQP7, AQP8 and AQP 11. Overall, current findings indicate a major role of AQP8 in water influx and efltux for sperm volume regulation, which is required for natural fertilization. The preliminary data suggestive of a role for AQP7 in sperm glycerol metabolism needs further substantiation. The association of AQP 11 with the residual cytoplasm of elongated spermatids and the distal tail of spermatozoa supports the hypothesis of more than just a role in conferring water permeability and also in the turnover and recycling of surplus cellular components made redundant during spermiogenesis and spermiation. This would be crucial for the maintenance of a germinal epithelium functioning efficiently in the production of spermatozoa.