scholarly journals Cargo-mediated recruitment of the endocytic adaptor protein Sla1 in S. cerevisiae

2020 ◽  
Vol 133 (19) ◽  
pp. jcs247684
Author(s):  
Thomas O. Tolsma ◽  
Hallie P. Febvre ◽  
Deanna M. Olson ◽  
Santiago M. Di Pietro
Keyword(s):  
Plasma Membrane ◽  
Membrane Proteins ◽  
Point Mutations ◽  
Adaptor Protein ◽  
Adaptor Proteins ◽  
Coat Proteins ◽  
Plasma Membrane Proteins ◽  
Membrane Recruitment ◽  
Surface Localization ◽  
Protein Recruitment

ABSTRACTEndocytosis of plasma membrane proteins is mediated by their interaction with adaptor proteins. Conversely, emerging evidence suggests that adaptor protein recruitment to the plasma membrane may depend on binding to endocytic cargo. To test this idea, we analyzed the yeast adaptor protein Sla1, which binds membrane proteins harboring the endocytic signal NPFxD via the Sla1 SHD1 domain. Consistently, SHD1 domain point mutations that disrupted NPFxD binding caused a proportional reduction in Sla1–GFP recruitment to endocytic sites. Furthermore, simultaneous SHD1 domain point mutation and deletion of the C-terminal LxxQxTG repeat (SR) region linking Sla1 to coat proteins Pan1 and End3 resulted in total loss of Sla1–GFP recruitment to the plasma membrane. These data suggest that multiple interactions are needed for recruitment of Sla1 to the membrane. Interestingly, a Sla1 fragment containing just the third SH3 domain, which binds ubiquitin, and the SHD1 domain displayed broad surface localization, suggesting plasma membrane recruitment is mediated by interaction with both NPFxD-containing and ubiquitylated plasma membrane proteins. Our results also imply that a Sla1 NPF motif adjacent to the SR region might regulate the Sla1–cargo interaction, mechanistically linking Sla1 cargo binding to endocytic site recruitment.

scholarly journals High temporal resolution reveals simultaneous plasma membrane recruitment of the TPLATE complex subunits

2020 ◽  
Author(s):  
Jie Wang ◽  
Evelien Mylle ◽  
Alexander Johnson ◽  
Nienke Besbrugge ◽  
Geert De Jaeger ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Temporal Resolution ◽  
Live Cell Imaging ◽  
Adaptor Protein ◽  
High Temporal Resolution ◽  
Spatiotemporal Resolution ◽  
Membrane Recruitment ◽  
Localization Analysis ◽  
Dynamic Assembly ◽  
Protein Recruitment

AbstractThe TPLATE complex (TPC) is a key endocytic adaptor protein complex in plants. TPC contains six evolutionary conserved subunits and two plant specific subunits, AtEH1/Pan1 and AtEH2/Pan1, which are not associated with the hexameric subcomplex in the cytoplasm. To investigate the dynamic assembly of the octameric TPC at the plasma membrane (PM), we performed state-of-the-art dual-color live cell imaging at physiological and a lowered temperature. Our data show that lowering the temperature slows down endocytosis and thereby enhances the temporal resolution of the differential recruitment of endocytic components. Under both normal and lowered temperature conditions, the core TPC subunit TPLATE, and the AtEH/Pan1 proteins, exhibited simultaneous recruitment at the PM. These results, together with our co-localization analysis of different TPC subunits, allow us to conclude that in plant cells, TPC is not recruited to the PM sequentially but as an octameric complex.One sentence summaryLowering the temperature increases spatiotemporal resolution of protein recruitment at the plasma membrane.

scholarly journals Ubiquitination, quality control and degradation of membrane proteins – chance for therapies?

2018 ◽  
Vol 72 ◽  
pp. 512-525
Author(s):  
Donata Wawrzycka ◽  
Katarzyna Mizio
Keyword(s):  
Quality Control ◽  
Plasma Membrane ◽  
Membrane Proteins ◽  
Membrane Protein ◽  
Membrane Integrity ◽  
Adaptor Proteins ◽  
Control Mechanisms ◽  
Plasma Membrane Proteins ◽  
Yeast Saccharomyces Cerevisiae ◽  
Protein Ubiquitination

Plasma membrane integrity maintenance is crucial for cell survival. Plasma membrane proteins are under tight regulation and under certain conditions are actively removed from the membrane allowing cells to adapt to changing environment. Proteins blocked in the cell membrane may interact with other molecules and form toxic aggregates. Ubiquitin is one of the most important modifiers targeting proteins for degradation and/or regulating protein functions. Several quality control mechanisms have been identified in eukaryotic cells: chaperone- dependent system that recognizes and ubiquitinates misfolded or redundant membrane proteins; protein-intrinsic LID-degron system, based on recognition of degron and ARTs-Rsp5 network that controls quality of membrane transporters. Rsp5, a Nedd4-family E3 ubiquitin ligase, is crucial for plasma membrane proteins ubiquitination. Rsp5-dependent ubiquitin action acts as a sorting signal for internalization of a membrane protein via endocytosis, recognition by the ESCRT system and vacuolar degradation. Rsp5 builds poliUb-chains on K63 and recognizes substrates through various adaptor proteins. Most of the identified Rsp5 adaptors belongs to the α-arrestin family. Plasma membrane protein ubiquitination and degradation disorders may cause neurodegenerative and cardiovascular diseases. The yeast Saccharomyces cerevisiae is one of the best models for studying trafficking pathways of membrane proteins and ubiquitination systems.

Detergent fractionation with subsequent subtractive suppression hybridization as a tool for identifying genes coding for plasma membrane proteins

2009 ◽  
Vol 18 (6) ◽  
pp. 527-535 ◽  
Author(s):  
Andreas Lange ◽  
Claudia Kistler ◽  
Tanja B. Jutzi ◽  
Alexandr V. Bazhin ◽  
Claus Detlev Klemke ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Membrane Proteins ◽  
Plasma Membrane Proteins ◽  
Subtractive Suppression Hybridization

scholarly journals pH-dependent Cargo Sorting from the Golgi

2011 ◽  
Vol 286 (12) ◽  
pp. 10058-10065 ◽  
Author(s):  
Chunjuan Huang ◽  
Amy Chang
Keyword(s):  
Plasma Membrane ◽  
Membrane Proteins ◽  
Atpase Activity ◽  
Secretory Pathway ◽  
Ph Dependence ◽  
Glucose Deprivation ◽  
Ph Homeostasis ◽  
Plasma Membrane Proteins ◽  
Cytosolic Ph ◽  
Cargo Sorting

The vacuolar proton-translocating ATPase (V-ATPase) plays a major role in organelle acidification and works together with other ion transporters to maintain pH homeostasis in eukaryotic cells. We analyzed a requirement for V-ATPase activity in protein trafficking in the yeast secretory pathway. Deficiency of V-ATPase activity caused by subunit deletion or glucose deprivation results in missorting of newly synthesized plasma membrane proteins Pma1 and Can1 directly from the Golgi to the vacuole. Vacuolar mislocalization of Pma1 is dependent on Gga adaptors although no Pma1 ubiquitination was detected. Proper cell surface targeting of Pma1 was rescued in V-ATPase-deficient cells by increasing the pH of the medium, suggesting that missorting is the result of aberrant cytosolic pH. In addition to mislocalization of the plasma membrane proteins, Golgi membrane proteins Kex2 and Vrg4 are also missorted to the vacuole upon loss of V-ATPase activity. Because the missorted cargos have distinct trafficking routes, we suggest a pH dependence for multiple cargo sorting events at the Golgi.

scholarly journals 5-Iodonaphthyl-1-azide labeling of plasma membrane proteins adjacent to specific sites via energy transfer

1997 ◽  
Vol 1324 (2) ◽  
pp. 320-332 ◽  
Author(s):  
Bruce I Meiklejohn ◽  
Noorulhuda A Rahman ◽  
Deborah A Roess ◽  
B.George Barisas
Keyword(s):  
Plasma Membrane ◽  
Energy Transfer ◽  
Membrane Proteins ◽  
Plasma Membrane Proteins

scholarly journals Role of the Endocytic Machinery in the Sorting of Lysosome-associated Membrane Proteins

2005 ◽  
Vol 16 (9) ◽  
pp. 4231-4242 ◽  
Author(s):  
Katy Janvier ◽  
Juan S. Bonifacino
Keyword(s):  
Plasma Membrane ◽  
Coat Protein ◽  
Membrane Proteins ◽  
Adaptor Protein ◽  
The Other ◽  
Sorting Signals ◽  
Efficient Delivery ◽  
Endocytic Machinery

The limiting membrane of the lysosome contains a group of transmembrane glycoproteins named lysosome-associated membrane proteins (Lamps). These proteins are targeted to lysosomes by virtue of tyrosine-based sorting signals in their cytosolic tails. Four adaptor protein (AP) complexes, AP-1, AP-2, AP-3, and AP-4, interact with such signals and are therefore candidates for mediating sorting of the Lamps to lysosomes. However, the role of these complexes and of the coat protein, clathrin, in sorting of the Lamps in vivo has either not been addressed or remains controversial. We have used RNA interference to show that AP-2 and clathrin—and to a lesser extent the other AP complexes—are required for efficient delivery of the Lamps to lysosomes. Because AP-2 is exclusively associated with plasma membrane clathrin coats, our observations imply that a significant population of Lamps traffic via the plasma membrane en route to lysosomes.

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