Vacuolar sorting receptors （VSRs） are type I integral membrane proteins that mediate the vacuolar transport of soluble cargo proteins via prevacuolar compartments （PVCs） in plants. Confocal immunofluorescent and i...Vacuolar sorting receptors （VSRs） are type I integral membrane proteins that mediate the vacuolar transport of soluble cargo proteins via prevacuolar compartments （PVCs） in plants. Confocal immunofluorescent and immunogold Electron Microscope （EM） studies have localized VSRs to PVCs or multivesicular bodies （MVBs） and trans-Golgi network （TGN） in various plant cell types, including suspension culture cells, root cells, developing and germinating seeds. Here, we provide evidence that VSRs reach plasma membrane （PM） in growing pollen tubes. Both immunofluorescent and immunogold EM studies with specific VSR antibodies show that, in addition to the previously demonstrated PVC/MVB localization, VSRs also localize to PM in lily and tobacco pollen tubes prepared from chemical fixation or high-pressure freezing/frozen substitution. Such a PM localization suggests an additional role of VSR proteins in mediating protein transport to PM and endocytosis in growing pollen tubes. Using a high-speed Spinning Disc Confocal Microscope, the possible fusion between VSR-positive PVC organelles and the PM was also observed in living tobacco pollen tubes transiently expressing the PVC reporter GFP-VSR. In contrast, the lack of a prominent PM localization of GFP-VSR in living pollen tubes may be due to the highly dynamic situation of vesicular transport in this fast-growing cell type.展开更多
Dear Editor,Over the last 15 years endocytosis has moved from being a process of only minor importance to plant physiologists to being one of the most exciting research areas in plant cell biology. These days, nobody ...Dear Editor,Over the last 15 years endocytosis has moved from being a process of only minor importance to plant physiologists to being one of the most exciting research areas in plant cell biology. These days, nobody doubts the operation of clathrin-mediated endocytosis as a mechanism for the internalization of a range of physiologically important transmembrane protein complexes at the plasma membrane （PM） of plant cells. These include both receptors and transporters. As in animal cells, most of these proteins are constitutively recycled back to the PM from an early endosome （EE）. However, some are destined for degradation and proceed further downstream in the endocytic pathway to late endosomes （LE）, where they are internalized into the intraluminal vesicles of the LE. Fusion of the LE with the lysosome/vacuole releases the vesicles leading to their degradation. The signal that marks PM proteins for degradation has been known in mammalian and yeast cells for quite some time and is polyubiquitination （Mukhopadhyay and Riezman, 2007）. In their Spotlight article, Zelazny and Vert （2015） highlight recent publications from the plant field that also demonstrate a key role for multiple monoubiquitination in the endocytosis of the metal transporters IRT1 （Barberon et al., 2011）, BOR1 （Kasai et al., 2011）, as well as lysine63-1inked polyubiquitination in vacuolar sorting of the auxin transporter PIN2 （Leitner et al., 2012）. They also draw attention to the discovery of a ubiquitin-binding protein （FREE1 ; also termed FYVE1 by Barberon et al., 2014） which locates to the LE in plant cells and is part of the ESCRT-I （endosomal sorting complex for transport） complex （Gao et al., 2014）. This complex sequesters ubiquitinated membrane cargo proteins and internalizes them. Finally, Zelazny and Vert draw attention to the work of Ivanov et al. （2014） in showing an increase in IRT1 degradation and iron deficiency in SNX1 mutants. So far so good, but Zelazny and Vert go on to conclude that a portion of the internalized IRT1 molecules that reach the LE are recycled back to the EE, a process that they consider to be mediated by sorting nexin 1 （SNX1）, a retromer protein. I question the soundness of this scenario.展开更多
文摘Vacuolar sorting receptors （VSRs） are type I integral membrane proteins that mediate the vacuolar transport of soluble cargo proteins via prevacuolar compartments （PVCs） in plants. Confocal immunofluorescent and immunogold Electron Microscope （EM） studies have localized VSRs to PVCs or multivesicular bodies （MVBs） and trans-Golgi network （TGN） in various plant cell types, including suspension culture cells, root cells, developing and germinating seeds. Here, we provide evidence that VSRs reach plasma membrane （PM） in growing pollen tubes. Both immunofluorescent and immunogold EM studies with specific VSR antibodies show that, in addition to the previously demonstrated PVC/MVB localization, VSRs also localize to PM in lily and tobacco pollen tubes prepared from chemical fixation or high-pressure freezing/frozen substitution. Such a PM localization suggests an additional role of VSR proteins in mediating protein transport to PM and endocytosis in growing pollen tubes. Using a high-speed Spinning Disc Confocal Microscope, the possible fusion between VSR-positive PVC organelles and the PM was also observed in living tobacco pollen tubes transiently expressing the PVC reporter GFP-VSR. In contrast, the lack of a prominent PM localization of GFP-VSR in living pollen tubes may be due to the highly dynamic situation of vesicular transport in this fast-growing cell type.
文摘Dear Editor,Over the last 15 years endocytosis has moved from being a process of only minor importance to plant physiologists to being one of the most exciting research areas in plant cell biology. These days, nobody doubts the operation of clathrin-mediated endocytosis as a mechanism for the internalization of a range of physiologically important transmembrane protein complexes at the plasma membrane （PM） of plant cells. These include both receptors and transporters. As in animal cells, most of these proteins are constitutively recycled back to the PM from an early endosome （EE）. However, some are destined for degradation and proceed further downstream in the endocytic pathway to late endosomes （LE）, where they are internalized into the intraluminal vesicles of the LE. Fusion of the LE with the lysosome/vacuole releases the vesicles leading to their degradation. The signal that marks PM proteins for degradation has been known in mammalian and yeast cells for quite some time and is polyubiquitination （Mukhopadhyay and Riezman, 2007）. In their Spotlight article, Zelazny and Vert （2015） highlight recent publications from the plant field that also demonstrate a key role for multiple monoubiquitination in the endocytosis of the metal transporters IRT1 （Barberon et al., 2011）, BOR1 （Kasai et al., 2011）, as well as lysine63-1inked polyubiquitination in vacuolar sorting of the auxin transporter PIN2 （Leitner et al., 2012）. They also draw attention to the discovery of a ubiquitin-binding protein （FREE1 ; also termed FYVE1 by Barberon et al., 2014） which locates to the LE in plant cells and is part of the ESCRT-I （endosomal sorting complex for transport） complex （Gao et al., 2014）. This complex sequesters ubiquitinated membrane cargo proteins and internalizes them. Finally, Zelazny and Vert draw attention to the work of Ivanov et al. （2014） in showing an increase in IRT1 degradation and iron deficiency in SNX1 mutants. So far so good, but Zelazny and Vert go on to conclude that a portion of the internalized IRT1 molecules that reach the LE are recycled back to the EE, a process that they consider to be mediated by sorting nexin 1 （SNX1）, a retromer protein. I question the soundness of this scenario.
