Biology subjects

Romao, L.

Publications and source records attributed to Romao, L..

5 recordsLinked to original sources

bioRxiv2023

Δ246p53 is a new p53 isoform that responds to DNA damage and regulates tumour growth

p53 is with little doubt one of the most powerful genes in our genome, as it makes growth vs arrest, repair vs replacement, catabolism vs anabolism, life vs death decisions in the cell. An alteration or malfunction in p53 may lead to cancer, degeneration or premature ageing. So, it is not surprising that p53 is also one of the most complex and tightly regulated genes, encoding for at least 10 RNA variants and 12 widely accepted protein forms. Here we identify one new p53 protein isoform of approximately 18 kDa that we termed {Delta}246p53. {Delta}246p53 is translated from an alternative translation initiation site (TIS) in codon 246. TIS-246 is preceded by a strong Kozak sequence and appears conserved in vertebrates, from sea lamprey to humans. {Delta}246p53s origin and expression in cells were confirmed by frameshift and start codon mutations; five different antibodies against several epitopes from its N-terminus down to its C-terminus and one against the DNA binding domain region just upstream of its initiator methionine working as negative control; as well as siRNAs and an antisense oligo targeting TIS-246, which knocked-down {Delta}246p53 with little or no effect on full-length (FL) p53 protein levels. {Delta}246p53 was induced by DNA damage and triggered senescence and impaired tumour formation/growth in colony formation assays. Mechanistically, {Delta}246p53 interacted with FLp53, leading to a decrease in the expression of downstream target genes HDM2 and p21. Concurrently, it also interacted with {Delta}Np63, leading to p53-independent p63-mediated activation of p21, a known senescence activator. Our results unveil a new naturally occurring and tightly controlled factor with specific functions in p21 regulation and senescence. Future studies on {Delta}246p53 are likely to help us better understand and control the processes of tumour suppression and ageing.

cancer biology↗

bioRxiv2023

Internal Translation of p53 Oncoproteins During Integrated Stress Response Confers Survival Advantage on Cancer Cells

p53 is the most known and studied tumour suppressor gene. Yet we have recently shown that p53 is also a proto-oncogene, as it encodes the {Delta}160p53 oncoprotein. Integrated stress response (ISR) is a survival pathway frequently activated in cancers, marked by the phosphorylation of eukaryotic initiation factor 2alpha (eIF2) and a defined reprogramming in mRNA translation. Here we identified ISR as a powerful trigger of p53 oncogene, leading to the induction of not only {Delta}160p53 but also {Delta}133p53, another protein variant of the p53 gene. Upon ISR the two isoforms were translated internally from p53 full-length (FL) transcript through an internal regulator of expression site (IRES) located in the vicinity of codon 160. Frameshift mutations upstream of codons 133 and 160 demonstrated that FLp53 protein synthesis is not required for making {Delta}133p53 and {Delta}160p53. Instead, targeting IRES(160) with an antisense oligo was sufficient to efficiently and specifically impair the expression of these isoforms without affecting FLp53 levels. This in turn averted ISRs protective program culminating in cancer cell cycle arrest and death. Mechanistically, FLp53 showed 3 times more affinity to {Delta}160p53 than to other isoforms, {Delta}40p53 or {Delta}133p53. During ISR {Delta}160p53 localized to the nucleus and strongly inhibited FLp53-mediated activation of pro-apoptotic gene p53 upregulated modulator of apoptosis (PUMA). Our results uncover a new branch of the ISR network essential for cancer cell survival and growth and establish the proof of concept for a new strategy to target cancer.

cancer biology↗

bioRxiv2022

SARS-CoV-2 spike protein induces long-term TLR4-mediated synapse and cognitive loss recapitulating Post-COVID syndrome

Cognitive dysfunction is often reported in post-COVID patients, but its underlying mechanisms remain unknown. While some evidence indicate that SARS-CoV-2 can reach and directly impact the brain, others suggest viral neuroinvasion as a rare event. Independently of brain viral infection, the ability of SARS-CoV-2 spike (S) protein to cross the BBB and reach memory-related brain regions has already been shown. Here, we demonstrate that brain infusion of S protein in mice induces late cognitive impairment and increases serum levels of neurofilament light chain (NFL), which recapitulates post-COVID features. Neuroinflammation, hippocampal microgliosis and synapse loss are induced by S protein. Increased engulfment of hippocampal presynaptic terminals late after S protein brain infusion were found to temporally correlate with cognitive deficit in mice. Blockage of TLR4 signaling prevented S-associated detrimental effects on synapse and memory loss. In a cohort of 86 patients recovered from mild COVID-19, genotype GG TLR4 -2604G>A (rs10759931) was associated with poor cognitive outcome. Collectively, these findings indicate that S protein directly impacts the brain and suggest that TLR4 is a potential target to prevent post-COVID cognitive dysfunction. One Sentence SummaryTLR4 mediates long-term cognitive impairment in mice and its genetic variant increases the risk of poor cognitive outcome in post-COVID patients.

neuroscience↗

bioRxiv2020

Membrane Elastic Properties During Neural Progenitor/Neural Stem Cell Differentiation

Neural stem cells differentiate into several cell types that display distinct functions. However, little is known about how cell surface mechanics vary during the differentiation process. Here, by precisely measuring membrane tension and bending modulus, we map their variations and correlate them with changes in cell morphology along differentiation into neurons, astrocytes and oligodendrocytes. Both neurons and undifferentiated cells reveal a decrease in membrane tension over the first hours of differentiation followed by stabilization, with no change in bending modulus. Astrocytes membrane tension initially decreases and then increases after 72h, accompanied by consolidation of GFAP expression and striking actin reorganization, while bending modulus increases following observed alterations. For oligodendrocytes, the changes in membrane tension are less abrupt over the first hours but their values subsequently decrease, correlating with a shift from O4 to MBP expressions and a remarkable actin reorganization, while bending modulus remains constant. Oligodendrocytes at later differentiation stages show membrane vesicles with similar membrane tension but higher bending modulus as compared to the cell surface. Altogether, our results display an entire spectrum of how membrane elastic properties are varying, contributing to a better understanding of neural stem cell differentiation from a mechanobiological perspective.

biophysics↗

bioRxiv2019

A role for DIS3L2 over human nonsense-mediated mRNA decay targets

The nonsense-mediated decay (NMD) pathway selectively degrades mRNAs carrying a premature translation-termination codon but also regulates the abundance of a large number of physiological mRNAs that encode full-length proteins. In human cells, NMD-targeted mRNAs are degraded by endonucleolytic cleavage and exonucleolytic degradation from both 5 and 3 ends. This is done by a process not yet completely understood that recruits decapping and 5-to-3 exonuclease activities, as well as deadenylating and 3-to-5 exonuclease exosome activities. In yeast, DIS3/Rrp44 protein is the catalytic subunit of the exosome, but in humans, there are three known paralogues of this enzyme: DIS3, DIS3L1, and DIS3L2. DIS3L1 and DIS3L2 exoribonucleases localize in the same compartment where NMD occurs, but little is known about their role in this process. In order to unveil the role of DIS3L2 in NMD, here we show that some NMD-targets accumulate in DIS3L2-depleted cells. mRNA half-life analysis further supports that these NMD-targets are in fact DIS3L2 substrates. Besides, we observed that DIS3L2 acts over full-length transcripts, through a process that also involves UPF1. Moreover, DIS3L2-mediated decay is dependent on the activity of the terminal uridylyl transferases Zcchc6/11 (TUT7/4). Together, our findings establish a role for DIS3L2 and uridylation in NMD.

molecular biology↗