It is still continuously debated whether the low-dose/dose-rate (LDR) of ionizing radiation represents a hazard for humans. Model organisms, such as fruit flies, are considered valuable systems to reveal insights into this issue. We found that, in wild-type Drosophila melanogaster larval neuroblasts, the frequency of Chromosome Breaks (CBs), induced by acute γ-irradiation, is considerably reduced when flies are previously exposed to a protracted dose of 0.4 Gy delivered at a dose rate of 2.5 mGy/h. This indicates that this exposure, which is associated with an increased expression of DNA damage response proteins, induces a radioadaptive response (RAR) that protects Drosophila from extensive DNA damage. Interestingly, the same exposure reduces the frequency of telomere fusions (TFs) from Drosophila telomere capping mutants suggesting that the LDR can generally promote a protective response on chromatin sites that are recognized as DNA breaks. Deep RNA sequencing revealed that RAR is associated with a reduced expression of Loquacious D (Loqs-RD) gene that encodes a well-conserved dsRNA binding protein required for esiRNAs biogenesis. Remarkably, loss of Loqs mimics the LDR-mediated chromosome protection as it decreases the IR-induced CBs and TFs frequency. Thus, our molecular characterization of RAR identifies Loqs as a key factor in the cellular response to LDR and in the epigenetic routes involved in radioresistance.

Chromobox Protein 3 (CBX3) overexpression is a common event occurring in cancer, promotes cancer cell proliferation and represents a poor prognosis marker in a plethora of human cancers. Here we describe that a wide spectrum of human cancers harbors a co-amplification of CBX3 gene with either EGFR or RAC1, which yields a statistically significant increase of both mRNA and protein levels of CBX3, EGFR and RAC1. We also reveal that the simultaneous overexpression of CBX3, RAC1 and EGFR gene products correlates with a worse prognosis compared to the condition when CBX3, RAC1 and EGFR are singularly upregulated. Furthermore, we also show that a co-occurrence of low-grade amplification, in addition to high-grade amplification, between CBX3 and EGFR or RAC1 is associated with a reduced patient lifespan. Finally, we find that CBX3 and RAC1/EGFR genetically interact in the model organism Drosophila melanogaster, suggesting that the simultaneous overexpression as well as well the co-occurrence of high- or low-grade copy number alterations in these genes is not accidental and could reflect evolutionarily conserved functional relationships.

Separase is a well conserved endopeptidase that facilitates sister chromatid separation at the metaphase-anaphase transition by cleaving cohesins. Beyond its role in chromosome segregation, Separase also participates in various biological processes, including chromatin organization and replication, centrosome disengagement and duplication, cytokinesis, and telomere capping. Here, we report that the loss of Drosophila separase ( Sse ) function induces significant changes in global protein expression and affects the protein levels of both A/C-type lamin C ( LamC ) and B-type lamin Dm0 ( Dm0 ). We further demonstrate that SSE physically interacts with lamins and colocalizes with them at the nuclear envelope during interphase. Additionally, loss of SSE activity disrupts nuclear organization in larval muscles and impairs locomotion in adult flies. Notably, similar to SSE in flies, depletion of human separase ( ESPL1 ) in SV40 fibroblasts leads to misshapen nuclei and increased levels of lamin A. Moreover, we show that ESPL1 interacts with lamin A in human fibroblasts, suggesting that the functional interaction between Separase and lamins is evolutionarily conserved across different organisms.

Compelling evidence indicates that defects in nucleocytoplasmic transport contribute to the pathogenesis of amyotrophic lateral sclerosis (ALS). In particular, hexanucleotide (G4C2) repeat expansions in C9orf72, the most common cause of genetic ALS, have a widespread impact on the transport machinery that regulates the nucleocytoplasmic distribution of proteins and RNAs. We previously reported that the expression of G4C2 hexanucleotide repeats in cultured human and mouse cells caused a marked accumulation of poly(A) mRNAs in the cell nuclei. To further characterize the process, we set out to systematically identify the specific mRNAs that are altered in their nucleocytoplasmic distribution in the presence of C9orf72-ALS RNA repeats. Interestingly, pathway analysis showed that the mRNAs involved in membrane trafficking are particularly enriched among the identified mRNAs. Most importantly, functional studies in cultured cells and Drosophila indicated that C9orf72 toxic species affect the membrane trafficking route regulated by ADP-Ribosylation Factor 1 GTPase Activating Protein (ArfGAP-1), which exerts its GTPase-activating function on the small GTPase ADP-ribosylation factor 1 to dissociate coat proteins from Golgi-derived vesicles. We demonstrate that the function of ArfGAP-1 is specifically affected by expanded C9orf72 RNA repeats, as well as by C9orf72-related dipeptide repeat proteins (C9-DPRs), indicating the retrograde Golgi-to-ER vesicle-mediated transport as a target of C9orf72 toxicity.

Separase plays a central role in chromosome separation during mitosis and in centrosome cycle. Tight control of separase activity is required to prevent unscheduled resolution of sister chromatid cohesion and centrosome aberrations, thereby preserving genome stability. In mammals, despite their disassembly in early mitosis, some nuclear envelope components possess mitotic roles, but links with separase activity remain unexplored. Here, we uncover a new mechanism of separase regulation involving lamin B1, a key nuclear envelope factor. We show that separase and lamin B1 associate preferentially during early stages of mitosis. Importantly, lamin B1 depletion leads to an increase in separase recruitment on chromosomes together with premature chromatid separation, a phenotype reminiscent of separase overexpression. Conversely, similar to separase depletion, lamin B1 overexpression induces formation of diplochromosomes- resulting from chromatid separation failure-, in association with centrosome amplification. Importantly, increasing separase level prevents lamin B1-induced centrosome aberrations, suggesting a separase defect at their origin. Indeed, we show that overexpression of lamin B1 leads to a decrease in the recruitment of separase to the chromosome and a delay in its activity. Taken together, this study unveils a novel mechanism of separase regulation involving the nuclear envelope factor lamin B1, that is crucial for genome integrity maintenance.