PAR-5 is a PARty hub in the germline

As our understanding of how molecular machineries work expands, an increasing number of proteins that appear as regulators of different processes have been identified. These proteins are hubs within and among functional networks. The 14-3-3 protein family is involved in multiple cellular pathways and, therefore, influences signaling in several disease processes, from neurobiological disorders to cancer. As a consequence, 14-3-3 proteins are currently being investigated as therapeutic targets. Moreover, 14-3-3 protein levels have been associated with resistance to chemotherapies. There are seven 14-3-3 genes in humans, while Caenorhabditis elegans only possesses two, namely par-5 and ftt-2. Among the C. elegans scientific community, par-5 is mainly recognized as one of the par genes that is essential for the asymmetric first cell division in the embryo. However, a recent study from our laboratory describes roles of par-5 in germ cell proliferation and in the cellular response to DNA damage induced by genotoxic agents. In this review, we explore the broad functionality of 14-3-3 proteins in C. elegans and comment on the potential use of worms for launching a drugs/modifiers discovery platform for the therapeutic regulation of 14-3-3 function in cancer

Modeling of autosomal-dominant retinitis pigmentosa in Caenorhabditis elegans uncovers a nexus between global impaired functioning of certain splicing factors and cell type-specific apoptosis

Retinitis pigmentosa (RP) is a rare genetic disease that causes gradual blindness through retinal degeneration. Intriguingly, seven of the 24 genes identified as responsible for the autosomal-dominant form (adRP) are ubiquitous spliceosome components whose impairment causes disease only in the retina. The fact that these proteins are essential in all organisms hampers genetic, genomic, and physiological studies, but we addressed these difficulties by using RNAi in Caenorhabditis elegans. Our study of worm phenotypes produced by RNAi of splicing-related adRP (s-adRP) genes functionally distinguishes between components of U4 and U5 snRNP complexes, because knockdown of U5 proteins produces a stronger phenotype. RNA-seq analyses of worms where s-adRP genes were partially inactivated by RNAi, revealed mild intron retention in developing animals but not in adults, suggesting a positive correlation between intron retention and transcriptional activity. interestingly, RNAi of s-adRP genes produces an increase in the expression of atl-1 (homolog of human ATR), which is normally activated in response to replicative stress and certain DNA-damaging agents. The up-regulation of atl-1 correlates with the ectopic expression of the pro-apoptotic gene egl-1 and apoptosis in hypodermal cells, which produce the cuticle, but not in other cell types. Our model in C. elegans resembles s-adRP in two aspects: The phenotype caused by global knockdown of s-adRP genes is cell type-specific and associated with high transcriptional activity. Finally, along with a reduced production of mature transcripts, we propose a model in which the retina-specific cell death in s-adRP patients can be induced through genomic instability

RSR-2, the Caenorhabditis elegans Ortholog of Human Spliceosomal Component SRm300/SRRM2, Regulates Development by Influencing the Transcriptional Machinery

Protein components of the spliceosome are highly conserved in eukaryotes and can influence several steps of the gene expression process. RSR-2, the Caenorhabditis elegans ortholog of the human spliceosomal protein SRm300/SRRM2, is essential for viability, in contrast to the yeast ortholog Cwc21p. We took advantage of mutants and RNA interference (RNAi) to study rsr-2 functions in C. elegans, and through genetic epistasis analysis found that rsr-2 is within the germline sex determination pathway. Intriguingly, transcriptome analyses of rsr-2(RNAi) animals did not reveal appreciable splicing defects but instead a slight global decrease in transcript levels. We further investigated this effect in transcription and observed that RSR-2 colocalizes with DNA in germline nuclei and coprecipitates with chromatin, displaying a ChIP-Seq profile similar to that obtained for the RNA Polymerase II (RNAPII). Consistent with a novel transcription function we demonstrate that the recruitment of RSR-2 to chromatin is splicing-independent and that RSR-2 interacts with RNAPII and affects RNAPII phosphorylation states. Proteomic analyses identified proteins associated with RSR-2 that are involved in different gene expression steps, including RNA metabolism and transcription with PRP-8 and PRP-19 being the strongest interacting partners. PRP-8 is a core component of the spliceosome and PRP-19 is the core component of the PRP19 complex, which interacts with RNAPII and is necessary for full transcriptional activity. Taken together, our study proposes that RSR-2 is a multifunctional protein whose role in transcription influences C. elegans development

A Histone Methylation Network Regulates Transgenerational Epigenetic Memory in C. elegans

How epigenetic information is transmitted from generation to generation remains largely unknown. Deletion of the C. elegans Histone H3 lysine 4 dimethyl (H3K4me2) demethylase spr-5 leads to inherited accumulation of the euchromatic H3K4me2 mark and progressive decline in fertility. Here we identified multiple chromatin-modifying factors, including novel H3K4me1/me2 and H3K9me3 methyltransferases, an H3K9me3 demethylase and an H3K9me reader, which either suppress or accelerate the progressive transgenerational phenotypes of spr-5 mutant worms. Our findings uncover a network of chromatin regulators that control the trans-generational flow of epigenetic information, and suggest that the balance between euchromatic H3K4 and heterochromatic H3K9 methylation regulates trans-generational effects on fertility

Cell cycle checkpoints in Caenorhabditis elegans: the 14-3-3 gene par-5 is required for germline development and DNA damage response / Checkpoints del ciclo celular en Caenorhabditis elegans: el gen 14-3-3, par-5, es necesario para el desarrollo y respuesta al daño genómico de la línea germinal

[eng] 14-3-3 proteins have been extensively studied from yeast to mammals, and are associated with multiple roles ranging from fundamental processes such as cell cycle, apoptosis and stress response to diseases such as neurodegeneration and cancer. Indeed, 14-3-3 proteins have been suggested as possible therapeutic targets in cancer treatment. There are seven 14-3-3 genes in mammals, whereas there are only two in Caenorhabditis elegans, ftt-2 and par-5. The ftt-2 gene is expressed only in somatic lineages, whereas par-5 expression is detected in both soma and germline. Although it is known that par-5 inactivation results in sterility, the role of this gene in germline development is poorly characterized. In the present study, we use a par-5 mutation and RNA interference to characterize par-5 functions in the germline. The lack of par-5 in germ cells causes cell cycle deregulation, the accumulation of endogenous DNA damage and genomic instability. Moreover, par-5 is required for checkpoint-induced cell cycle arrest in response to DNA-damaging agents. We propose a model whereby PAR-5 regulates CDK-1 phosphorylation to prevent premature mitotic entry. Even though mammalian 14-3-3 homologs have diverged into seven genes, we verified that the basic functions of 14-3-3 in cell cycle control have been conserved in C. elegans. Therefore, this study opens a new path to investigate molecular mechanisms of 14-3-3 proteins and establishes C. elegans as a suitable system to screen for genes (RNAi libraries or mutagenesis), and drugs which can modify 14-3-3 functions.[spa] Las proteínas 14-3-3 han sido ampliamente estudiadas desde levadura hasta mamíferos y han sido asociadas con múltiples roles en procesos como ciclo celular, apoptosis y la respuesta al estrés. Así mismo estas proteínas se han visto involucradas en enfermedades neurodegenerativas y cáncer. De hecho, las proteínas 14-3-3 han sido propuestas como posibles agentes terapéuticos en el tratamiento contra el cáncer. En mamíferos existen 7 genes que codifican para proteínas 14-3-3, mientras en Caenorhabditis elegans solo hay dos, ftt-2 and par-5. El gen ftt-2 sólo es expresado en células somáticas, mientras par-5 se expresa tanto en células somáticas como en la línea germinal. Aunque se sabe que la inactivación de par-5 puede producir esterilidad, el rol de este gen en el desarrollo de la línea germinal no ha sido caracterizado. En este estudio, se usó una mutación de par-5 y RNA interferente para caracterizar la función de par-5 en la línea germinal. La falta de par-5 en la línea germinal causa una desregulación del ciclo celular, acumulación de daño genómico e inestabilidad genómica. Además, par-5 es requerido para el arresto celular inducido por el checkpoint en respuesta a los agentes que dañan el genoma. A partir de los resultados obtenidos, se propone un modelo según el cual PAR-5 regula la fosforilación de CDK-1 para prevenir la entrada prematura en mitosis. Aunque los homólogos de 14-3-3 en humanos han divergido en 7 genes, este estudio permitió verificar que las funciones básicas de las proteínas 14-3-3 en el control ciclo celular están conservadas en C. elegans. Por lo tanto, este estudio abre un nuevo camino para estudiar las funciones moleculares de las proteínas 14-3-3 y establece C. elegans como un modelo adecuado para la búsqueda de genes y/o drogas que modifiquen la función de las proteínas 14-3-3

Developmental dynamics of gene expression and alternative polyadenylation in the Caenorhabditis elegans germline

Abstract Background The 3′ untranslated regions (UTRs) of mRNAs play a major role in post-transcriptional regulation of gene expression. Selection of transcript cleavage and polyadenylation sites is a dynamic process that produces multiple transcript isoforms for the same gene within and across different cell types. Using LITE-Seq, a new quantitative method to capture transcript 3′ ends expressed in vivo, we have characterized sex- and cell type-specific transcriptome-wide changes in gene expression and 3′UTR diversity in Caenorhabditis elegans germline cells undergoing proliferation and differentiation. Results We show that nearly half of germline transcripts are alternatively polyadenylated, that differential regulation of endogenous 3′UTR variants is common, and that alternative isoforms direct distinct spatiotemporal protein expression patterns in vivo. Dynamic expression profiling also reveals temporal regulation of X-linked gene expression, selective stabilization of transcripts, and strong evidence for a novel developmental program that promotes nucleolar dissolution in oocytes. We show that the RNA-binding protein NCL-1/Brat is a posttranscriptional regulator of numerous ribosome-related transcripts that acts through specific U-rich binding motifs to down-regulate mRNAs encoding ribosomal protein subunits, rRNA processing factors, and tRNA synthetases. Conclusions These results highlight the pervasive nature and functional potential of patterned gene and isoform expression during early animal development

RSR-2, the Caenorhabditis elegans Ortholog of Human Spliceosomal Component SRm300/SRRM2, Regulates Development by Influencing the Transcriptional Machinery

Protein components of the spliceosome are highly conserved in eukaryotes and can influence several steps of the gene expression process. RSR-2, the Caenorhabditis elegans ortholog of the human spliceosomal protein SRm300/SRRM2, is essential for viability, in contrast to the yeast ortholog Cwc21p. We took advantage of mutants and RNA interference (RNAi) to study rsr-2 functions in C. elegans, and through genetic epistasis analysis found that rsr-2 is within the germline sex determination pathway. Intriguingly, transcriptome analyses of rsr-2(RNAi) animals did not reveal appreciable splicing defects but instead a slight global decrease in transcript levels. We further investigated this effect in transcription and observed that RSR-2 colocalizes with DNA in germline nuclei and coprecipitates with chromatin, displaying a ChIP-Seq profile similar to that obtained for the RNA Polymerase II (RNAPII). Consistent with a novel transcription function we demonstrate that the recruitment of RSR-2 to chromatin is splicing-independent and that RSR-2 interacts with RNAPII and affects RNAPII phosphorylation states. Proteomic analyses identified proteins associated with RSR-2 that are involved in different gene expression steps, including RNA metabolism and transcription with PRP-8 and PRP-19 being the strongest interacting partners. PRP-8 is a core component of the spliceosome and PRP-19 is the core component of the PRP19 complex, which interacts with RNAPII and is necessary for full transcriptional activity. Taken together, our study proposes that RSR-2 is a multifunctional protein whose role in transcription influences C. elegans development

Modeling of autosomal-dominant retinitis pigmentosa in Caenorhabditis elegans uncovers a nexus between global impaired functioning of certain splicing factors and cell type-specific apoptosis

Retinitis pigmentosa (RP) is a rare genetic disease that causes gradual blindness through retinal degeneration. Intriguingly, seven of the 24 genes identified as responsible for the autosomal-dominant form (adRP) are ubiquitous spliceosome components whose impairment causes disease only in the retina. The fact that these proteins are essential in all organisms hampers genetic, genomic, and physiological studies, but we addressed these difficulties by using RNAi in Caenorhabditis elegans. Our study of worm phenotypes produced by RNAi of splicing-related adRP (s-adRP) genes functionally distinguishes between components of U4 and U5 snRNP complexes, because knockdown of U5 proteins produces a stronger phenotype. RNA-seq analyses of worms where s-adRP genes were partially inactivated by RNAi, revealed mild intron retention in developing animals but not in adults, suggesting a positive correlation between intron retention and transcriptional activity. interestingly, RNAi of s-adRP genes produces an increase in the expression of atl-1 (homolog of human ATR), which is normally activated in response to replicative stress and certain DNA-damaging agents. The up-regulation of atl-1 correlates with the ectopic expression of the pro-apoptotic gene egl-1 and apoptosis in hypodermal cells, which produce the cuticle, but not in other cell types. Our model in C. elegans resembles s-adRP in two aspects: The phenotype caused by global knockdown of s-adRP genes is cell type-specific and associated with high transcriptional activity. Finally, along with a reduced production of mature transcripts, we propose a model in which the retina-specific cell death in s-adRP patients can be induced through genomic instability

RSR-2, the <i>Caenorhabditis elegans</i> Ortholog of Human Spliceosomal Component SRm300/SRRM2, Regulates Development by Influencing the Transcriptional Machinery

<div><p>Protein components of the spliceosome are highly conserved in eukaryotes and can influence several steps of the gene expression process. RSR-2, the <i>Caenorhabditis elegans</i> ortholog of the human spliceosomal protein SRm300/SRRM2, is essential for viability, in contrast to the yeast ortholog Cwc21p. We took advantage of mutants and RNA interference (RNAi) to study <i>rsr-2</i> functions in <i>C. elegans</i>, and through genetic epistasis analysis found that <i>rsr-2</i> is within the germline sex determination pathway. Intriguingly, transcriptome analyses of <i>rsr-2(RNAi)</i> animals did not reveal appreciable splicing defects but instead a slight global decrease in transcript levels. We further investigated this effect in transcription and observed that RSR-2 colocalizes with DNA in germline nuclei and coprecipitates with chromatin, displaying a ChIP-Seq profile similar to that obtained for the RNA Polymerase II (RNAPII). Consistent with a novel transcription function we demonstrate that the recruitment of RSR-2 to chromatin is splicing-independent and that RSR-2 interacts with RNAPII and affects RNAPII phosphorylation states. Proteomic analyses identified proteins associated with RSR-2 that are involved in different gene expression steps, including RNA metabolism and transcription with PRP-8 and PRP-19 being the strongest interacting partners. PRP-8 is a core component of the spliceosome and PRP-19 is the core component of the PRP19 complex, which interacts with RNAPII and is necessary for full transcriptional activity. Taken together, our study proposes that RSR-2 is a multifunctional protein whose role in transcription influences <i>C. elegans</i> development.</p></div