Plasma Membrane Protein Nce102 Modulates Morphology and Function of the Yeast Vacuole

Abstract

Membrane proteins are targeted not only to specific membranes in the cell architecture, but also to distinct lateral microdomains within individual membranes to properly execute their biological functions. Yeast tetraspan protein Nce102 has been shown to migrate between such microdomains within the plasma membrane in response to an acute drop in sphingolipid levels. Combining microscopy and biochemistry methods, we show that upon gradual ageing of a yeast culture, when sphingolipid demand increases, Nce102 migrates from the plasma membrane to the vacuole. Instead of being targeted for degradation it localizes to V-ATPase-poor, i.e., ergosterol-enriched, domains of the vacuolar membrane, analogous to its plasma membrane localization. We discovered that, together with its homologue Fhn1, Nce102 modulates vacuolar morphology, dynamics, and physiology. Specifically, the fusing of vacuoles, accompanying a switch of fermenting yeast culture to respiration, is retarded in the strain missing both proteins. Furthermore, the absence of either causes an enlargement of ergosterol-rich vacuolar membrane domains, while the vacuoles themselves become smaller. Our results clearly show decreased stability of the V-ATPase in the absence of either Nce102 or Fhn1, a possible result of the disruption of normal microdomain morphology of the vacuolar membrane. Therefore, the functionality of the vacuole as a whole might be compromised in these cells.

Touch and Go: Membrane Contact Sites Between Lipid Droplets and Other Organelles

“…Vph1, a component of vacuolar proton pump ATPase, is excluded from L o microdomains. In contrast, sterol transporters (LaM6/Ltn1 and Nce102), TORC1 (target of rapamycin complex 1) subunits (Tco89, Tor Complex I 89) and subunits or interactors of the TORC1-regulating EGO/ragulator complex (Ivy1, Interacting with Vps33 and Ypt7; Gtr1 and 2, GTP binding protein resemblance 1 and 2; Iml1, increased minichromosome loss 1) are enriched in L o microdomains ( Toulmay and Prinz, 2013 ; Wang et al, 2014 ; Murley et al, 2015 , 2017 ; Numrich et al, 2015 ; Varlakhanova et al, 2018 ; Vaskovicova et al, 2020 ).…”

Lipid Droplets and Their Autophagic Turnover via the Raft-Like Vacuolar Microdomains

“…Remarkably, all 15 vacuolar membrane proteins tested in this study segregated between the two microdomains, with the majority of them being localized to the liquid-disordered domain and only two proteins, Gtr2 and Ivy1p, sitting in the liquid-ordered domain that resembles lipid rafts in the plasma membrane. Further studies added Atg6 and Atg14 (next section), Lam6/Ltc1, Gtr1, Iml1, Tco89, and Nce102 (see below) to the list of raft-like vacuolar domain-associated proteins [ 33 , 130 , 131 , 132 ].…”

“…Another raft-like vacuolar domain-associated protein that regulates the formation of this domain is Nce102 [ 132 ]. This protein partially re-localizes from the lipid rafts in the plasma membrane to the raft-like microdomains in the vacuolar membrane via endocytosis and the multivesicular body pathway when cells enter the stationary phase.…”

“…This protein partially re-localizes from the lipid rafts in the plasma membrane to the raft-like microdomains in the vacuolar membrane via endocytosis and the multivesicular body pathway when cells enter the stationary phase. Interestingly, Nce102 negatively regulates the size of the liquid-ordered domains because in the nce102 mutant, they are substantially larger correlating with the increased intravacuolar degradation of the liquid-disordered domain marker, Vph1 [ 132 ]. Regarding other non-Atg raft-like domain-associated proteins, they were not implicated in microdomain formation.…”

The phosphatase Glc7 controls eisosomal response to starvation via posttranslational modification of Pil1

“…A screen for PI(4,5)P2 regulators revealed eisosome factors Slm1 and Slm2 also bind lipids, which are required for proper eisosomal organisation and integrate with TORC2 signalling and lipid synthesis (Audhya et al, 2004; Berchtold et al, 2012; Kamble et al, 2011; Riggi et al, 2018). Other examples of proteins required for proper eisosome assembly include Nce102, a sphingolipid sensor (Fröhlich et al, 2009; Vaskovicova et al, 2020; Zahumenský et al, 2022), and Seg1, a stability factor that operates upstream of Pil1 (Moreira et al, 2012; Seger et al, 2011).…”