Biology subjects

Portela, P.

Publications and source records attributed to Portela, P..

4 recordsLinked to original sources

bioRxiv2026

A Genome-wide Visual Screen Identifies Lysophosphatidylcholine as Counter Spatial Regulator of DAG and Sterols in Yeast

Membrane lipids are heterogeneously distributed across the bilayers of cellular membranes. Cytosolic-facing pools of diacylglycerol (DAG) in the yeast Saccharomyces cerevisiae are enriched at both ends of the endomembrane system from the vacuolar membrane to the polarized plasma membrane (PM) of buds. However, how this distribution is maintained remains unknown. Using a genome-wide DAG biosensor screen in yeast, we identify regulators of DAG spatial distribution, enriched in proteins involved in vesicle or lipid transport and in phospholipid or sterol metabolism. A subset of mutants exhibited DAG mislocalization predominantly to the PM, with the most severe phenotype linked to a mutant of a predicted lipase we have named Drl1 (DAG redistribution lipase 1). Reversion of this phenotype required both enzymatic activity and the presence of an intrinsically disordered carboxy-terminal domain. Lipidomic analysis revealed that loss of Drl1 increased cellular lysophosphatidylcholine (LysoPC) levels. Remarkably, we find that supplementing cells with a non-metabolizable LysoPC analogue replicated the mutant DAG phenotype, implicating LysoPC as a novel spatial regulator of DAG. High-resolution imaging suggests that LysoPC reduces the PM sterol pool resulting in DAG expansion into new PM territories. More globally, our work expands the known interplay between various lipids and their co-regulation to maintain accurate membrane properties.

cell biology↗

bioRxiv2021

Heat Stress Regulates the Expression of TPK1 Gene at Transcriptional and Post-Transcriptional Levels in Saccharomyces cerevisiae

In Saccharomyces cerevisiae, cAMP regulates a number of different cellular processes, such as cell growth, metabolism, stress resistance and gene transcription. The intracellular target for this second messenger in yeast cells is the cAMP-dependent protein kinase (PKA). The way in which a broad specificity protein kinase mediates one right physiological response after cAMP increase indicates that specificity is highly regulated in the cAMP / PKA system. Here we address the mechanism through which cAMP-PKA signalling mediates its response to heat shock thermotolerance in Saccharomyces cerevisiae. Yeast PKA is a tetrameric holoenzyme composed of a regulatory (Bcy1) subunit dimer and two catalytic subunits (Tpk1, Tpk2 and Tpk3). PKA subunits are differentially expressed under certain stress conditions. In the present study we show that, although the mRNA levels of TPK1 are upregulated upon heat shock at 37{degrees}C, no change is detected in Tpk1 protein levels. The half-life of TPK1 mRNA increases and this mRNA condensates in cytoplasmic foci upon thermal stress. The resistance of TPK1 mRNA foci to cycloheximide-induced disassembly, together with the polysome profiling analysis suggest that TPK1 mRNA is impaired for entry into translation. TPK1 mRNA foci and TPK1 expression were also evaluated during thermotolerance. The crosstalk of cAMP-PKA pathway and cell wall integrity (CWI) signalling was also studied. Wsc3 sensor and other components of the CWI pathway are necessary for the upregulation of TPK1 mRNA upon heat shock conditions. The assembly in cytoplasmic foci upon thermal stress shows to be dependent of Wsc3. Finally, evidence of an increase in the abundance of Tpk1 in the PKA holoenzyme in response to heat shock is presented, suggesting that a recurrent stress enhanced the fitness for the coming favorable conditions The results indicate the existence of a mechanism that exclusively regulates Tpk1 subunit expression and therefore contributing to the specificity of cAMP-PKA. SUMMARY STATEMENTPKA subunits are differentially expressed under heat-shock conditions. The mRNA of the TPK1 subunit is upregulated upon heat-shock at 37{degrees}C and thermotolerance, the half-life increases upon heat-stress and also this transcript condensates in cytoplasmic foci upon thermal stress and thermotolerance. The resistance to cycloheximide treatment of TPK1 mRNA foci together with the analysis by polysome profiling suggest that TPK1 mRNA is impaired for entry into translation upon thermal stress. An increase in Tpk1 protein and PKA activity was detected after the heat stress treatments. Cell Wall Integrity pathway, through Wsc3 sensor, is involved in TPK1 expression. Heat-stress regulates TPK1 expression through this pathway from an intermediate step of the cascade and independently of the upstream elements of the CWI pathway activation. These results demonstrate a new crosstalk between the two signalling pathways. The increment in Tpk1-dependent PKA activity during cell adaptation to heat stress might contribute to the overall cellular fitness when more favorable environmental conditions are restored. The results indicate the existence of a mechanism that exclusively regulates TPK1 subunit expression and therefore contributing to the specificity of cAMP-PKA.

molecular biology↗

bioRxiv2019

CoFe granules focus coordinated glycolytic mRNA localization and translation to fuel glucose fermentation

Glycolysis is a fundamental metabolic pathway for glucose catabolism across biology, and glycolytic enzymes are amongst the most abundant proteins in cells. Their expression at such levels provides a particular challenge. Here we demonstrate that the glycolytic mRNAs are localized to granules in yeast and human cells. Detailed live cell and smFISH studies in yeast show that the mRNAs are actively translated in granules, and this translation appears critical for the localization. Furthermore, this arrangement is likely to facilitate the higher level organisation and control of the glycolytic pathway. Indeed, the degree of fermentation required by cells is intrinsically connected to the extent of mRNA localization to granules. On this basis, we term these granules, Core Fermentation (CoFe) granules; they appear to represent translation factories allowing high-level co-ordinated enzyme synthesis for a critical metabolic pathway.

cell biology↗

bioRxiv2018

Translation factor mRNA granules direct protein synthetic capacity to regions of polarized growth

mRNA localization serves key functions in localized protein production making it critical that the translation machinery itself is present at these locations. Here we show that translation factor mRNAs are localized to distinct granules within yeast cells. In contrast to many mRNP granules, such as P-bodies and stress granules, which contain translationally repressed mRNAs, these granules harbor translated mRNAs under active growth conditions. The granules require Pablp for their integrity and are inherited by developing daughter cells in a She2p/ She3p dependent manner. These results point to a model where roughly half the mRNA for certain translation factors are specifically directed in granules toward the tip of the developing daughter cell where protein synthesis is most heavily required, which has particular implications for filamentous forms of growth. Such a feedforward mechanism would ensure adequate provision of the translation machinery where it is to be needed most over the coming growth cycle. SummaryThis study shows that mRNAs encoding a range of translation factors are localized to granules that get transported into the yeast daughter cell using the She2p/She3p machinery. This likely supports an intensification of protein synthetic activity to facilitate apical polarized growth.

cell biology↗