John calarco

Alternative pre-mRNA splicing has the potential to greatly diversify the repertoire of transcripts in multicellular organisms. Increasing evidence suggests that this expansive layer of gene regulation plays a particularly important role in the development and function of the nervous system, one of the most complex organ systems found in nature. In this review, we highlight recent studies that continue to emphasize the influence and contribution of alternative splicing regulation to various aspects of neuronal development in addition to its role in the mature nervous system...

Alternative splicing plays a key role in the expansion of proteomic and regulatory complexity, yet the functions of the vast majority of differentially spliced exons are not known. In this study, we observe that brain and other tissue-regulated exons are significantly enriched in flexible regions of proteins that likely form conserved interaction surfaces. These proteins participate in significantly more interactions in protein-protein interaction (PPI) networks than other proteins. Using LUMIER, an automated PPI assay, we observe that approximately one-third of analyzed neural-regulated exons affect PPIs...

The adult mammalian central nervous system exhibits restricted regenerative potential. Chen et al. (2011) and El Bejjani and Hammarlund (2012) used Caenorhabditis elegans to uncover intrinsic factors that inhibit regeneration of axotomized mature neurons, opening avenues for potential therapeutics.

Recent genome-wide analyses have indicated that almost all primary transcripts from multi-exon human genes undergo alternative pre-mRNA splicing (AS). Given the prevalence of AS and its importance in expanding proteomic complexity, a major challenge that lies ahead is to determine the functional specificity of isoforms in a cellular context. A significant fraction of alternatively spliced transcripts are regulated in a tissue- or cell-type-specific manner, suggesting that these mRNA variants likely function in the generation of cellular diversity...

Alternative splicing (AS) plays a crucial role in the diversification of gene function and regulation. Consequently, the systematic identification and characterization of temporally regulated splice variants is of critical importance to understanding animal development. We have used high-throughput RNA sequencing and microarray profiling to analyze AS in C. elegans across various stages of development. This analysis identified thousands of novel splicing events, including hundreds of developmentally regulated AS events...

Alternative splicing has a crucial role in the generation of biological complexity, and its misregulation is often involved in human disease. Here we describe the assembly of a 'splicing code', which uses combinations of hundreds of RNA features to predict tissue-dependent changes in alternative splicing for thousands of exons. The code determines new classes of splicing patterns, identifies distinct regulatory programs in different tissues, and identifies mutation-verified regulatory sequences. Widespread regulatory strategies are revealed, including the use of unexpectedly large combinations of features, the establishment of low exon inclusion levels that are overcome by features in specific tissues, the appearance of features deeper into introns than previously appreciated, and the modulation of splice variant levels by transcript structure characteristics...

Cwc21 (complexed with Cef1 protein 21) is a 135 amino acid yeast protein that shares homology with the N-terminal domain of human SRm300/SRRM2, a large serine/arginine-repeat protein shown previously to associate with the splicing coactivator and 3'-end processing stimulatory factor, SRm160. Proteomic analysis of spliceosomal complexes has suggested a role for Cwc21 and SRm300 at the core of the spliceosome. However, specific functions for these proteins have remained elusive. In this report, we employ quantitative genetic interaction mapping, mass spectrometry of tandem affinity-purified complexes, and microarray profiling to investigate genetic, physical, and functional interactions involving Cwc21...

Alternative splicing is a key process underlying the evolution of increased proteomic and functional complexity and is especially prevalent in the mammalian nervous system. However, the factors and mechanisms governing nervous system-specific alternative splicing are not well understood. Through a genome-wide computational and expression profiling strategy, we have identified a tissue- and vertebrate-restricted Ser/Arg (SR) repeat splicing factor, the neural-specific SR-related protein of 100 kDa (nSR100). We show that nSR100 regulates an extensive network of brain-specific alternative exons enriched in genes that function in neural cell differentiation...

RATIONALE: Fragile histidine triad (FHIT) is a tumor suppressor gene involved in the pathogenesis of lung cancer. OBJECTIVES: The purpose of this study was to investigate the different molecular alterations leading to the inactivation of FHIT gene function and to validate their use as biomarkers of risk for progression of the disease in patients belonging to the multicentric European study for the Early detection of Lung Cancer (EUELC) who were resected for early-stage lung tumors...

During the past approximately 20 years, studies on alternative splicing (AS) have largely been directed at the identification and characterization of factors and mecha nisms responsible for the control of splice site selection, using model substrates and on a case by case basis. These studies have provided a wealth of information on the factors and interactions that control formation of the spliceosome. However, relatively little is known about the global regulatory properties of AS. Important questions that need to be addressed are: which exons are alternatively spliced and under which cellular contexts, what are the functional roles of AS events in different cellular contexts, and how are AS events controlled and coordinated with each other and with other levels of gene regulation to achieve cell- and development-specific functions...

Alternative splicing is a powerful mechanism affording extensive proteomic and regulatory diversity from a limited repertoire of genes. However, the extent to which alternative splicing has contributed to the evolution of primate species-specific characteristics has not been assessed previously. Using comparative genomics and quantitative microarray profiling, we performed the first global analysis of alternative splicing differences between humans and chimpanzees. Surprisingly, 6%-8% of profiled orthologous exons display pronounced splicing level differences in the corresponding tissues from the two species...

Communication between neurons is largely achieved through chemical synapses, where neurotransmitters are released from synaptic vesicles at presynaptic terminals to activate postsynaptic cells. Exo- and endocytosis are coordinated to replenish the synaptic vesicle pool for sustained neuronal activity. We identified syd-9 (syd, synapse defective), a gene that encodes multiple C2H2 zinc-finger domain-containing proteins specifically required for synaptic function in Caenorhabditis elegans. syd-9 loss-of-function mutants exhibit locomotory defects, a diffuse distribution of synaptic proteins, and decreased synaptic transmission with unaffected neurodevelopment...

Spinal muscular atrophy is a neurodegenerative disorder caused by the deletion or mutation of the survival-of-motor-neuron gene, SMN1. An SMN1 paralog, SMN2, differs by a C-->T transition in exon 7 that causes substantial skipping of this exon, such that SMN2 expresses only low levels of functional protein. A better understanding of SMN splicing mechanisms should facilitate the development of drugs that increase survival motor neuron (SMN) protein levels by improving SMN2 exon 7 inclusion. In addition, exonic mutations that cause defective splicing give rise to many genetic diseases, and the SMN1/2 system is a useful paradigm for understanding exon-identity determinants and alternative-splicing mechanisms...

In a recent study, we provided evidence that strong promoter-bound transcriptional activators result in higher levels of splicing and 3'-end cleavage of nascent pre-mRNA than do weak promoter-bound activators and that this effect of strong activators requires the carboxyl-terminal domain (CTD) of RNA polymerase II (pol II). In the present study, we have investigated the mechanism of activator- and CTD-mediated stimulation of pre-mRNA processing. Affinity chromatography experiments reveal that two factors previously implicated in the coupling of transcription and pre-mRNA processing, PSF and p54(nrb)/NonO, preferentially bind a strong rather than a weak activation domain...