Biology subjects

Sepulveda, C.

Publications and source records attributed to Sepulveda, C..

3 recordsLinked to original sources

bioRxiv2024

BAX and BAK expression triggers multiple IRE1 signaling outputs under ER stress

Adaptation to endoplasmic reticulum (ER) stress depends on the activation of the unfolded protein response (UPR) stress sensor inositol-requiring enzyme 1 alpha (IRE1). IRE1 is a central ER stress sensors, that signals through the activation of its RNase domain to catalyze the splicing the mRNA encoding the transcription factor X-box binding protein 1 (XBP1), resulting on the expression of a stable and active transcription factor termed XBP1s. The kinetics and amplitude of IRE1 signaling are regulated by different posttranslational modifications and the physical interaction of different factors. Early studies demonstrated that the expression of the proapoptotic proteins BAX and BAX enhance UPR signaling. However, the possible effects on the RNase activity were not defined. Here we provide preliminary evidence indicating that BAX and BAK deficiency increases the in 10 folds the threshold of ER stress to induce XBP1 mRNA splicing, and the upregulation of its target genes. In addition, the degradation of RIDD substrates was strongly reduced in BAX and BAK null cells. BAX and BAK double deficiency also attenuated the levels of IRE1 phosphorylation under mild ER stress. These results reinforce previous findings indicating that proapoptotic BAX and BAK have alternative functions at the ER regulating the UPR.

cell biology↗

bioRxiv2021

A KARRIKIN INSENSITIVE2 paralog in lettuce mediates highly sensitive germination responses to karrikinolide

Karrikins (KARs) are chemicals in smoke that can enhance germination of many plants. Lactuca sativa cv. Grand Rapids (lettuce), germinates in the presence of nanomolar karrikinolide (KAR1). We found that lettuce is much less responsive to KAR2 or a mixture of synthetic strigolactone analogs, rac-GR24. We investigated the molecular basis of selective and sensitive KAR1 perception in lettuce. The lettuce genome contains two copies of KARRIKIN INSENSITIVE2 (KAI2), a receptor that is required for KAR responses in Arabidopsis thaliana. LsKAI2b is more highly expressed than LsKAI2a in dry achenes and during early stages of seed imbibition. Through cross-species complementation assays in Arabidopsis we found that LsKAI2b confers robust responses to KAR1, but LsKAI2a does not. Therefore, LsKAI2b likely mediates KAR1 responses in lettuce. We compared homology models of the ligand-binding pockets of KAI2 proteins from lettuce and a fire follower, Emmenanthe penduliflora. This identified pocket residues 96, 124, 139, and 161 as candidates that influence the ligand-specificity of KAI2. Further support for the significance of these residues was found through a broader comparison of pocket residue conservation among 324 asterid KAI2 proteins. We tested the effects of substitutions at these four positions in Arabidopsis thaliana KAI2 and found that a broad array of responses to KAR1, KAR2, and rac-GR24 could be achieved.

plant biology↗

bioRxiv2019

VPS41 recessive mutation causes ataxia and dystonia with retinal dystrophy and mental retardation by inhibiting HOPS function and mTORC1 signaling

The vacuolar protein sorting protein 41 (VPS41) is a neuroprotective protein in models of Parkinsons disease (PD). As part of the HOPS (Homotypic fusion and Protein Sorting) complex, VPS41 regulates fusion of lysosomes with late endosomes and autophagosomes. Independent of HOPS, VPS41 regulates transport of newly synthesized lysosomal membrane proteins and secretory proteins. Here we report two brothers with compound heterozygous mutations in VPS41 (VPS41R662* and VPS41S285P), born to healthy and non-consanguineous parents. Both patients displayed transient retinal dystrophy, ataxia and dystonia, with brain MRI findings of cerebellar atrophy and a thin saber-shape corpus callosum. Patient-derived fibroblasts contained enzymatically active lysosomes that were poorly reached by endocytic cargo and failed to attract the mTORC1 complex. Consequently, transcription factor TFE3, a driver of autophagy and lysosomal genes, showed continuous nuclear localization which resulted in elevated LC3-II levels and an impaired response to nutrient starvation. CRISPR/CAS VPS41 HeLa knockout cells showed a similar phenotype that could be rescued by wildtype VPS41 but not by VPS41S285P or VPS41R662*. mTORC1 inhibition was also seen after knockout of HOPS subunits VPS11 or VPS18. Regulated neuropeptide secretion in PC12 VPS41 knockout cells was rescued by VPS41S285P expression, indicating that this HOPS-independent function was preserved. Co-expression of the VPS41S285P and VPS41R662* variants in a C. elegans model of PD abolished the protective effect of VPS41 against -synuclein-induced neurodegeneration. We conclude that both disease-associated VPS41 variants specifically abrogate HOPS function, which leads to a delay in endocytic cargo delivery to lysosomes, mTORC1 inhibition and irresponsiveness to autophagic clues. Our studies signify a link between HOPS function and mTORC1 signaling and imply that HOPS function is required for the neuroprotective effect of VPS41 in PD.

cell biology↗