Biology subjects

Roiuk, M.

Publications and source records attributed to Roiuk, M..

5 recordsLinked to original sources

bioRxiv2026

Stress induces DCP2 translation via a stalling-dependent mechanism

The integrated stress response (ISR) globally suppresses protein synthesis while selectively permitting translation of a small subset of stress-responsive mRNAs, many of which contain upstream or overlapping open reading frames (uORFs/oORFs). Although translational induction of transcripts such as ATF4 has classically been attributed to delayed re-initiation caused by reduced ternary complex availability, the mechanisms by which uORFs and oORFs allow ISR-selective translation remain incompletely understood. Here, using ribosome profiling during early ISR activation combined with reporter assays, we identify DCP2, encoding a major mRNA decapping enzyme, as a previously unrecognized ISR-induced transcript. We show that translational induction of DCP2 depends on an overlapping ORF whose conserved 3' region, corresponding to a ribosome pausing site, acts as a potent inhibitory element. Both the DCP2 oORF and main ORF increase in translation during stress, indicating that stress relieves repression by this inhibitory element. This reveals a mode of ISR-dependent gene regulation in which inhibition by a nascent peptide or stalling element embedded either in a uORF or an oORF is relieved upon stress to induce translation.

molecular biology↗

bioRxiv2024

Human eIF2A has a minimal role in translation initiation and in uORF-mediated translational control

Initiation of translation on eukaryotic mRNAs requires a 40S ribosome loaded with an initiator tRNA in order to scan for, and to identify, an initiation codon. Under most conditions, the initiator tRNA is recruited to the ribosome as part of a ternary complex composed of initiator tRNA, eIF2 and GTP. Although this function of recruiting the initiator tRNA was originally ascribed to another factor, eIF2A, it was later disproven and shown to belong to eIF2. Nonetheless, eIF2A is still considered a translation initiation factor because it binds the ribosome and shows genetic interactions with other initiation factors such as eIF4E. The exact function of eIF2A during translation initiation, however, remains unclear. Here we systematically test in HeLa cells, using ribosome profiling and luciferase reporter assays, the role of eIF2A in translation initiation, including translation of upstream ORFs that are either initiated with a AUG or near-cognate codons. Since eIF2A is thought to take over the function of eIF2 when eIF2 is inhibited, we also test conditions where the integrated stress response is activated, thereby leading to eIF2 inactivation. In none of our assays, however, could we detect a role of eIF2A in translation initiation. It is possible that eIF2A plays a role in translation regulation in specific conditions that we have not tested here, or that it plays a role in a different aspect of RNA biology.

molecular biology↗

bioRxiv2024

Loss of the Familial Dysautonomia gene Elp1 in cerebellar granule cell progenitors leads to ataxia in mice

Familial Dysautonomia (FD) is an autosomal recessive disorder caused by a splice site mutation in the gene ELP1, which disproportionally affects neurons. While classically characterized by deficits in sensory and autonomic neurons, neuronal defects in the central nervous system have been described. ELP1 is highly expressed in the normal developing and adult cerebellum, but its role in cerebellum development is unknown. To investigate the cerebellar function of Elp1, we knocked out Elp1 in cerebellar granule cell progenitors (GCPs) and examined the outcome on animal behavior and cellular composition. We found that GCP-specific conditional knockout of Elp1 (Elp1cKO) resulted in ataxia by 8 weeks of age. Cellular characterization showed that the animals had smaller cerebella with fewer granule cells. This defect was already apparent 7 days after birth, when Elp1cKO animals also exhibited fewer mitotic GCPs and shorter Purkinje dendrites. Through molecular characterization, we found that loss of Elp1 was associated with an increase in apoptotic cell death and cell stress pathways in GCPs. Our study demonstrates the importance of ELP1 within the developing cerebellum, and suggests that Elp1 loss in the GC lineage may also play a role in the progressive ataxia phenotypes of FD patients.

neuroscience↗

bioRxiv2023

Niche signalling regulates eIF3d1 phosphorylation to promote distinct modes of translation initiation in stem and differentiating cells

Stem cells have the unique ability among adult cells to give rise to cells of different identities. To do so, they must change gene expression in response to environmental signals. Much work has focused on how transcription is regulated to achieve these changes, however in many cell types, transcripts and proteins correlate poorly, indicating that post-transcriptional regulation is important. To assess how translational control can influence stem cell fate, we use the Drosophila testis as a model. The testis niche secretes a ligand to activate the JAK/STAT pathway in two stem cell populations, germline stem cells (GSCs) and somatic cyst stem cells (CySCs). We find that global translation rates are high in CySCs and decrease during differentiation, and that JAK/STAT signalling regulates translation. To determine how translation was regulated, we knocked down translation initiation factors and found that the cap binding complex, eIF4F, is dispensable in differentiating cells, but is specifically required in CySCs for self-renewal, acting downstream of JAK/STAT activity. Moreover, we identify eIF3d1 as a key regulator of CySC fate, and show that its phosphorylation is critical to maintain CySC self-renewal. We further show that Casein Kinase II, which controls eIF3d1 phosphorylation, is sufficient to restore CySC function in the absence of JAK/STAT. We propose a model in which niche signals regulate a specific translation programme in which only some mRNAs are translated, through regulation of eIF3d phosphorylation. The mechanism we identify allows stem cells to switch between modes of translation, adding a layer of regulation on top of transcription and providing cells with the ability to rapidly change gene expression upon receiving external stimuli.

developmental biology↗

bioRxiv2022

PRRC2 proteins regulate translation initiation by promoting leaky scanning

Roughly half of animal mRNAs contain upstream Open Reading Frames (uORFs). These uORFs represent an impediment to translation of the main ORF since ribosomes usually bind the mRNA cap at the 5 end and then scan for ORFs in a 5-to-3 fashion. One way for ribosomes to bypass uORFs is via leaky scanning, whereby the ribosome disregards the uORF start codon. Hence leaky scanning is an important instance of post-transcriptional regulation that affects gene expression. Few molecular factors regulating or facilitating this process are known. Here we show that the PRRC2 proteins PRRC2A, PRRC2B and PRRC2C regulate translation initiation. We find that they bind eukaryotic translation initiation factors and preinitiation complexes, and are enriched on ribosomes translating mRNAs with uORFs. We find that PRRC2 proteins promote leaky scanning past translation start codons, thereby promoting translation of mRNAs containing uORFs. Since PRRC2 proteins have been associated with cancer, this provides a mechanistic starting point for understanding their physiological and pathophysiological roles.

molecular biology↗