Distinct endocytic pathways identified in tobacco pollen tubes using charged nanogold

Abstract

In an attempt to dissect endocytosis in Nicotiana tabacum L. pollen tubes, two different probes – positively or negatively charged nanogold – were employed. The destiny of internalized plasma membrane domains, carrying negatively or positively charged residues, was followed at the ultrastructural level and revealed distinct endocytic pathways. Time-course experiments and electron microscopy showed internalization of subapical plasma-membrane domains that were mainly recycled to the secretory pathway through the Golgi apparatus and a second mainly degradative pathway involving plasma membrane retrieval at the tip. In vivo time-lapse experiments using FM4-64 combined with quantitative analysis confirmed the existence of distinct internalization regions. Ikarugamycin, an inhibitor of clathrin-dependent endocytosis, allowed us to further dissect the endocytic process: electron microscopy and time-lapse studies suggested that clathrin-dependent endocytosis occurs in the tip and subapical regions, because recycling of positively charged nanogold to the Golgi bodies and the consignment of negatively charged nanogold to vacuoles were affected. However, intact positively charged-nanogold transport to vacuoles supports the idea that an endocytic pathway that does not require clathrin is also present in pollen tubes.

Pollen Proteins Bind to the C-terminal Domain of Nicotiana alata Pistil Arabinogalactan Proteins

“…For 120K to arrive at a vacuole, it must travel from the ECM through the pollen tube endomembrane system. Recent work has highlighted two distinct endocytic pathways in pollen tubes (54,55). Both clathrin-coated and noncoated vesicle endocytic pathways exist; clathrincoated vesicles traffic to the trans-Golgi, whereas noncoated vesicles travel to the multivescular body and a vacuole.…”

“…Both clathrin-coated and noncoated vesicle endocytic pathways exist; clathrincoated vesicles traffic to the trans-Golgi, whereas noncoated vesicles travel to the multivescular body and a vacuole. However, there is evidence that these two pathways are not mutually exclusive, suggesting that there is some degree of sorting of endocytic vesicles and their cargo (54). If endocytic vesicles contain many molecules with different destinations, these molecules need to be sorted consistently if they are to reliably arrive to their final destination.…”

A Membrane Microdomain-Associated Protein, Arabidopsis Flot1, Is Involved in a Clathrin-Independent Endocytic Pathway and Is Required for Seedling Development

“…Apart from the well-studied clathrin-dependent endocytosis (Chen et al, 2011), other entry pathways, including membrane raftmediated endocytosis, also operate in plant cells (Murphy et al, 2005;Moscatelli et al, 2007;Bandmann and Homann, 2011). Recent studies suggested that plant flotillins play a critical role in symbiotic bacterial infection of M. truncatula (Haney and Long, 2010;Haney et al, 2011), and the recycling of PIN proteins may depend on structural sterols and membrane rafts in Arabidopsis Willemsen et al, 2003;Men et al, 2008).…”

Exocytosis Precedes and Predicts the Increase in Growth in Oscillating Pollen Tubes

“…These vesicles move along actin microfilaments to the apical domain where they undergo exocytosis, fusing with the apical plasma membrane and contributing their contents to the cell wall (Heslop-Harrison, 1987;Steer and Steer, 1989;Lancelle and Hepler, 1992;Cheung and Wu, 2008). Due to the necessity of maintaining the appropriate surface-to-volume ratio, much of the secreted membrane is reclaimed by endocytosis (Picton and Steer, 1983;Moscatelli et al, 2007;Bove et al, 2008;Ketelaar et al, 2008). Endocytotic vesicles may quickly cycle back and revisit the apical plasma membrane, or they may join the reverse fountain streaming pattern and be transported toward the base of the pollen tube (Parton et al, 2001;Bove et al, 2008).…”