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Here we conduct a study involving 12 individuals with retinal dystrophy, neurological impairment, and skeletal abnormalities, with special focus on GPATCH11, a lesser-known G-patch domain-containing protein, regulator of RNA metabolism. To elucidate its role, we study fibroblasts from unaffected individuals and patients carrying the recurring c.328+1 G > T mutation, which specifically removes the main part of the G-patch domain while preserving the other domains. Additionally, we generate a mouse model replicating the patients’ phenotypic defects, including retinal dystrophy and behavioral abnormalities. Our results reveal a subcellular localization of GPATCH11 characterized by a diffuse presence in the nucleoplasm, as well as centrosomal localization, suggesting potential functions in RNA and cilia metabolism. Transcriptomic analysis performed on mouse retina detect dysregulation in both gene expression and splicing activity, impacting key processes such as photoreceptor light responses, RNA regulation, and primary cilia-associated metabolism. Proteomic analysis of mouse retina confirms the roles GPATCH11 plays in RNA processing, splicing, and transcription regulation, while also suggesting additional functions in synaptic plasticity and nuclear stress response. Our research provides insights into the diverse roles of GPATCH11 and identifies that the mutations affecting this protein are responsible for a recently characterized described syndrome. Here the authors identify GPATCH11 variants responsible for retinal dystrophy with neurological impairment and facial dysmorphy. They explore its function using a mouse model and demonstrate GPATCH11’s involvement in RNA regulation and splicing.

Genome replication is highly regulated in time and space, but the rules governing the remodeling of these programs during evolution remain largely unknown. We generated genome-wide replication timing profiles for ten Lachancea yeasts, covering a continuous evolutionary range from closely related to more divergent species. We show that replication programs primarily evolve through a highly dynamic evolutionary renewal of the cohort of active replication origins. We found that gained origins appear with low activity yet become more efficient and fire earlier as they evolutionarily age. By contrast, origins that are lost comprise the complete range of firing strength. Additionally, they preferentially occur in close vicinity to strong origins. Interestingly, despite high evolutionary turnover, active replication origins remain regularly spaced along chromosomes in all species, suggesting that origin distribution is optimized to limit large inter-origin intervals. We propose a model on the evolutionary birth, death, and conservation of active replication origins. Temporal programs of genome replication show different levels of conservation between closely or distantly related species. Here, the authors generate genome-wide replication timing profiles for ten yeast species, and analyze their evolutionary dynamics.

Physiological status can influence social behavior, which in turn can affect physiology and health. Previously, we reported that tumor growth in Drosophila virgin females depends on the social context, but did not investigate the underlying physiological mechanisms. Here, we sought to characterize the signal perceived between tumorous flies, ultimately discovering that the tumor suppressive effect varies depending on reproductive status. Firstly, we show that the tumor suppressive effect is neither dependent on remnant pheromone-like products nor on the microbiota. Transcriptome analysis of the heads of these tumorous flies reveals social-dependent gene-expression changes related to nervous-system activity, suggesting that a cognitive-like relay might mediate the tumor suppressive effect. The transcriptome also reveals changes in the expression of genes related to mating behavior. Surprisingly, we observed that this social-dependent tumor-suppressive effect is lost in fertilized females. After mating, Drosophila females change their behavior—favoring offspring survival—in response to peptides transferred via the male ejaculate, a phenomenon called “male manipulation”. Remarkably, the social-dependent tumor suppressive effect is restored in females mated by sex-peptide deficient males. Since male manipulation has likely been selected to favor male gene transmission, our findings indicate that this evolutionary trait impedes social-dependent tumor growth slowdown.

Clostridioides difficile is a major cause of nosocomial infections associated with antibiotic therapy classified as an urgent antibiotic resistance threat. This pathogen interacts with host and gut microbial communities during infection, but the mechanisms of these interactions remain largely to be uncovered. Noncoding RNAs contribute to bacterial virulence and host responses, but their expression has not been explored during C. difficile infection. We took advantage of the conventional mouse model of C. difficile infection to look simultaneously to the dynamics of gene expression in pathogen, its host, and gut microbiota composition, providing valuable resources for future studies. We identified a number of ncRNAs that could mediate the adaptation of C. difficile inside the host and the crosstalk with the host immune response. Promising inflammation markers and potential therapeutic targets emerged from this work open new directions for RNA-based and microbiota-modulatory strategies to improve the efficiency of C. difficile infection treatments.

Background Next-generation sequencing technologies have revolutionized the study of small RNAs (sRNAs) on a genome-wide scale. However, classical sRNA library preparation methods introduce serious bias, mainly during adapter ligation steps. Several types of sRNA including plant microRNAs (miRNA), piwi-interacting RNAs (piRNA) in insects, nematodes and mammals, and small interfering RNAs (siRNA) in insects and plants contain a 2’-O-methyl (2’-OMe) modification at their 3′ terminal nucleotide. This inhibits 3′ adapter ligation and makes library preparation particularly challenging. To reduce bias, the NEBNext kit (New England Biolabs) uses polyethylene glycol (PEG), the NEXTflex V2 kit (BIOO Scientific) uses both randomised adapters and PEG, and the novel SMARTer (Clontech) and CATS (Diagenode) kits avoid ligation altogether. Here we compared these methods with Illumina’s classical TruSeq protocol regarding the detection of normal and 2’ OMe RNAs. In addition, we modified the TruSeq and NEXTflex protocols to identify conditions that improve performance. Results Among the five kits tested with their respective standard protocols, the SMARTer and CATS kits had the lowest levels of bias but also had a strong formation of side products, and as a result performed relatively poorly with biological samples; NEXTflex detected the largest numbers of different miRNAs. The use of a novel type of randomised adapters called MidRand-Like (MRL) adapters and PEG improved the detection of 2’ OMe RNAs both in the TruSeq as well as in the NEXTflex protocol. Conclusions While it is commonly accepted that biases in sRNA library preparation protocols are mainly due to adapter ligation steps, the ligation-free protocols were not the best performing methods. Our modified versions of the TruSeq and NEXTflex protocols provide an improved tool for the study of 2’ OMe RNAs.

Small nucleolar RNAs are non-coding transcripts that guide chemical modifications of RNA substrates and modulate gene expression at the epigenetic and post-transcriptional levels. However, the extent of their regulatory potential and the underlying molecular mechanisms remain poorly understood. Here, we identify a conserved, previously unannotated intronic C/D-box snoRNA, termed snR107 , hosted in the fission yeast long non-coding RNA mamRNA and carrying two independent cellular functions. On the one hand, snR107 guides site-specific 25S rRNA 2’-O-methylation and promotes pre-rRNA processing and 60S subunit biogenesis. On the other hand, snR107 associates with the gametogenic RNA-binding proteins Mmi1 and Mei2, mediating their reciprocal inhibition and restricting meiotic gene expression during sexual differentiation. Both functions require distinct cis -motifs within snR107 , including a conserved 2’-O-methylation guiding sequence. Together, our results position snR107 as a dual regulator of rRNA modification and gametogenesis effectors, expanding our vision on the non-canonical functions exerted by snoRNAs in cell fate decisions. Small nucleolar RNAs (snoRNAs) are known for their role in RNA modification to regulate gene expression. Here, the authors identify a snoRNA that not only guides ribosomal RNA 2’- O-methylation but also modulates the activities of RNA-binding proteins involved in fission yeast gametogenesis.

The nucleoid-associated protein H-NS is a bacterial xenogeneic silencer responsible for preventing costly expression of genes acquired through horizontal gene transfer. H-NS silences several Salmonella Pathogenicity Islands (SPIs) essential for host infection. The stochastic expression of SPI-1 is required for invasion of host epithelial cells but complicates investigation of factors involved in SPI-1 chromatin structure and regulation. We performed functional genomics on sorted Salmonella populations expressing SPI-1 or not, to characterize how SPI-1 activation affects chromatin composition, DNA conformation, gene expression and SPI-1 subcellular localization. We show that silent SPIs are associated with spurious antisense transcriptional activity originating from H-NS-free regions. Upon SPI-1 activation, remodeling of H-NS occupancy defines a new chromatin landscape, which together with the master SPI-1 regulator HilD, facilitates transcription of SPI-1 genes. SPI-1 activation promotes formation of Transcription Induced Domains accompanied by repositioning SPI-1 close to the nucleoid periphery. We present a model for tightly regulated chromatin remodeling that minimizes the cost of pathogenicity island activation. The bacterial protein H-NS prevents costly expression of horizontally acquired genes such as those in Salmonella pathogenicity islands (SPIs), which are essential for infection. Here, Kortebi et al. show that the expression of SPI-1 is associated with Salmonella chromatin remodelling and with the repositioning of this region toward the nucleoid periphery.

We have cloned cDNAs for the human homologues of the yeast Dcp1 and Dcp2 factors involved in the major (5′–3′) and NMD mRNA decay pathways. While yeast Dcp1 has been reported to be the decapping enzyme, we show that recombinant human Dcp2 (hDcp2) is enzymatically active. Dcp2 activity appears evolutionarily conserved. Mutational and biochemical analyses indicate that the hDcp2 MutT/Nudix domain mediates this activity. hDcp2 generates m7GDP and 5′‐phosphorylated mRNAs that are 5′–3′ exonuclease substrates. Corresponding decay intermediates are present in human cells showing the relevance of this activity. hDcp1 and hDcp2 co‐localize in cell cytoplasm, consistent with a role in mRNA decay. Interestingly, these two proteins show a non‐uniform distribution, accumulating in specific foci.

In this work, by comparing genomes of closely related individuals of Streptomyces isolated at a spatial microscale (millimeters or centimeters), we investigated the extent and impact of horizontal gene transfer in the diversification of a natural Streptomyces population. We show that despite these conspecific strains sharing a recent common ancestor, all harbored significantly different gene contents , implying massive and rapid gene flux. The accessory genome of the strains was distributed across insertion/deletion events (indels) ranging from one to several hundreds of genes. Indels were preferentially located in the arms of the linear chromosomes (ca. 12 Mb) and appeared to form recombination hot spots. Some of them harbored biosynthetic gene clusters (BGCs) whose products confer an inhibitory capacity and may constitute public goods that can favor the cohesiveness of the bacterial population. Moreover, a significant proportion of these variable genes were either plasmid borne or harbored signatures of actinomycete integrative and conjugative elements (AICEs). We propose that conjugation is the main driver for the indel flux and diversity in Streptomyces populations. IMPORTANCE Horizontal gene transfer is a rapid and efficient way to diversify bacterial gene pools. Currently, little is known about this gene flux within natural soil populations. Using comparative genomics of Streptomyces strains belonging to the same species and isolated at microscale, we reveal frequent transfer of a significant fraction of the pangenome. We show that it occurs at a time scale enabling the population to diversify and to cope with its changing environment, notably, through the production of public goods.

Abstract The N6-methyladenosine (m6A) modification of HIV-1 has been widely studied but the number and precise positions of the m6A sites remain unclear due to the lack of precision of detection methods. Using the latest Nanopore chemistry and direct m6A base-calling, we identified 18 m6A: 14 at the 3′ end and 4 in central regions of the genome. Our data reveal differential methylation of these positions between splicing isoforms. Eleven of these sites are clustered in two short segments with peak-shaped methylation profiles. Single-molecule analysis revealed that a very small number of transcripts were unmethylated in both clusters. We also identified a ∼732 nt RNA species resulting from the transcription of non-integrated viral DNA circles closed by two long terminal repeats. These transcripts started in the first LTR, terminated at the polyA site of the second LTR, and harbored six m6A sites. Five of these sites were present in other transcripts and, remarkably, had the highest methylation rates. The sixth site was methylated only in this transcript, suggesting a role for this RNA in HIV-1 infection. These findings reveal a new landscape of HIV m6A transcriptome modifications and pave the way for studies deciphering their role in the viral life cycle.