Expression of Spliceosome-Associated Protein 49 Is Required for Early Embryogenesis inCaenorhabditis elegans

Fujita, Masaki; Kawano, Taizo; Terashima, Kazuhiro; Takai, Yoshimi; Sakamoto, Hiroshi

doi:10.1006/bbrc.1998.9747

Cited by 7 publications

(4 citation statements)

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“…There are no characterized mutations in core splicing factors in C. elegans, but the activities of some core splicing factors have been disrupted using the technique of RNA interference (11). These factors include both subunits of U2AF and the factor cSAP49, a subunit of the U2-associated factor SF3b (14,48). As expected, RNA interference of any of these three genes results in embryonic lethality.…”

Section: Discussionmentioning

confidence: 93%

Analysis of smu-1, a Gene That Regulates the Alternative Splicing of unc-52 Pre-mRNA in Caenorhabditis elegans

Spike

Shaw

Herman

2001

Molecular and Cellular Biology

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Mutations in the smu-1 gene of Caenorhabditis elegans were previously shown to suppress mutations in the genes mec-8 and unc-52. mec-8 encodes a putative RNA binding protein that affects the accumulation of specific alternatively spliced mRNA isoforms produced by unc-52 and other genes. unc-52 encodes a set of basement membrane proteins, homologs of mammalian perlecan, that are important for body wall muscle assembly and attachment to basement membrane, hypodermis, and cuticle. We show that a presumptive null mutation in smu-1 suppresses nonsense mutations in exon 17 but not exon 18 of unc-52 and enhances the phenotype conferred by an unc-52 splice site mutation in intron 16. We have used reverse transcription-PCR and RNase protection to show that loss-of-function smu-1 mutations enhance accumulation in larvae of an alternatively spliced isoform that skips exon 17 but not exon 18 of unc-52. We have identified smu-1 molecularly; it encodes a nuclearly localized protein that contains five WD motifs and is ubiquitously expressed. The SMU-1 amino acid sequence is more than 60% identical to a predicted human protein of unknown function. We propose that smu-1 encodes a trans-acting factor that regulates the alternative splicing of the pre-mRNA of unc-52 and other genes.Alternative splicing of pre-mRNA is a complex but common mechanism of regulating gene function in multicellular eukaryotes. Current estimates suggest that at least one-third of all genes in humans (17) are alternatively spliced. Patterns of alternative splicing are in some cases known to be cell type specific or developmentally regulated (16,24). Regulated alternative splicing of pre-mRNA most often results in the synthesis of multiple (functional) proteins from the same premRNA, but alternative splicing can also function as an on-off switch for gene function (1).Analysis of a genetic interaction in C. elegans has led to the proposal that the mec-8 gene encodes an RNA binding protein that regulates alternative splicing of unc-52 pre-mRNA (26). The unc-52 gene encodes, via alternative splicing and alternative 3Ј-end usage, homologs of perlecan, the core protein of the mammalian basement membrane heparan sulfate proteoglycan (39,40). A subset of UNC-52 isoforms is present in the basement membrane adjoining body wall muscle (12,30,33,39) and is important both for initiation of myofilament lattice assembly and for maintaining anchorages of body wall muscle to basement membrane, hypodermis, and cuticle. Null mutations in unc-52 lead to paralysis and arrest at the twofold stage of embryonic elongation (19,39,46); the muscle cells in these mutants have severely disorganized thick and thin filaments. Viable unc-52 mutants exhibit normal embryogenesis but suffer from a progressive disruption of body wall muscle and progressive paralysis during larval development (15). The unc-52 viable mutations are located in a region of unc-52 that when transcribed is alternatively spliced; most are nonsense mutations in alternatively spliced exons that prematurely truncate so...

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Section: Discussionmentioning

confidence: 93%

Analysis of smu-1, a Gene That Regulates the Alternative Splicing of unc-52 Pre-mRNA in Caenorhabditis elegans

Spike

Shaw

Herman

2001

Molecular and Cellular Biology

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“…Upregulation of another protein splicing factor 3B subunit 4 was only created by MEK1E and MKK5D. Previous studies indicated that the SAP49 protein was a component of the essential splicing factor SF3b and was associated with the U2 small nuclear RNA (Fujita et al, 1998;Bouck et al, 1998). The limited expression leads us to speculate that the phosphorus metabolism is involved in MEK1E, MKK3b and MKK5D function, while the alternative splicing of snRNA is needed for MEK1E and MKK5D function.…”

Section: Discussionmentioning

confidence: 93%

Proteomic approach to substrates related to MAPK pathway in 293T cells

Zhang

Wang

2007

Cell Biology International

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Abundant evidence indicates that potential scaffold proteins and adaptor or linker molecules organize and specify various MAP kinase cascades. In the present study, proteomic methodologies were applied to screen these potential molecules in combination with cell morphology and cell cycle analysis. MEK1E, MKK3b, MKK5D and MKK7D were selected as representative MKKs of four main MAPK pathways. Our results showed that similar morphological transformation and G(2)/M cell cycle arrest were promoted by the over-expressed four kinases. Furthermore, global change in response to the over-expressed four kinases was characterized by differential proteomics. Eleven distinctly changed proteins were detected, in which four proteins (serine/threonine kinase 4, glutathione S-transferase p1-1, glycoprotein IX and soluble inorganic pyrophosphatase) were reported to be relative to MAPK pathways, while the other seven proteins may be new elements of substrates of the kinases. In our experiment, the expression of platelet glycoprotein IX precursor, glutathione S-transferase p1-1, peroxiredoxin 6, Ras-related protein Rab-34 and arginase II, mitochondrial precursor was up-regulated, while the expression of serine/threonine kinase 4 (MST1) was down-regulated by the four kinases. These results suggest that these six proteins may be common targets of all the MAPK pathways in 293T cell line. Interestingly, the expression of splicing factor 3B subunit 4 and soluble inorganic pyrophosphatase (Ppase) was specifically up-regulated by MEK1E and MKK5D, and by MEK1E, MKK3b and MKK5D, respectively. The expression of methylglyoxalase was down-regulated by MEK1E and MKK7D. Furthermore, the expression of ADP-ribosylation factor-like protein 1 was up-regulated by MKK5D but down-regulated by MKK3b and MKK7D. These findings revealed the characteristic molecular responses to four MKKs. In conclusion, our study not only confirms that MST1, glutathione S-transferase p1-1, glycoprotein IX and soluble PPase belong to MAPK pathways, but also provides seven novel molecules for the further study of the pathways.

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“…The idea that RBM7 may be a splicing factor was initially based on its homology with other splicing factors, especially SAP49. Also known as splicing factor 3b sub‐unit 4, SAP49 is essential in the Caenorhabditis elegans worm (Fujita et al, 1998) and yeast ( Saccharomyces cerevisiae ) (Igel et al, 1998). It directly and stably interacts with SF3b subunit 2 (SAP145) and functions to tether U2 small nuclear ribonucleoprotein particle (snRNP) with pre‐mRNA at the branch site during spliceosome assembly (Champion‐Arnaud and Reed, 1994).…”

Section: Discussionmentioning

confidence: 99%

Spermatogenetic Expression of RNA‐Binding Motif Protein 7, a Protein That Interacts With Splicing Factors

Guo

Boros

Chan

et al. 2003

Journal of Andrology

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We have previously shown that a ubiquitously expressed RNA splicing factor, RNA-binding motif 7 (RBM7), cloned from a testis complementary DNA library, enhances messenger RNA (mRNA) splicing in vitro and is expressed in a cell-restricted fashion. Herein, we detail its mRNA and protein expression in the rodent testis. RNA in situ hybridization shows that Rbm7 expression in rat germ cells closely parallels the entry and progression of meiosis. The expression commences in type B spermatogonia, it rises during the preleptotene stage, peaks in leptotene spermatocytes, and declines afterward, but increases again in stage-associated pachytene spermatocytes. An affinity-purified polyclonal antibody raised against a peptide corresponding to amino acids 202-224 of the mouse RBM7 recognized the predicted 35 kd protein both in testicular lysates and in in vitro translation reactions. Consistent with the in situ hybridization results, RBM7 immunoreactivity was also detected in type B spermatogonia, spanned the entire period of spermatocyte development, and extended to round and early elongated spermatids. Moreover, RBM7 appeared nuclear up to the mid pachytene stage and became cytoplasmic thereafter. Consistent with its role in RNA splicing, yeast 2-hybrid and glutathione S-transferase pull-down assays show that RBM7 interacts with splicing factor 3b subunit 2 (SAP145), and with the splicing regulator, SRp20. These interactions and the nuclear localization of RBM7 provide insights into its function in pre-mRNA processing in developing spermatocytes during entry into meiosis and progression through the meiotic prophase.

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Expression of Spliceosome-Associated Protein 49 Is Required for Early Embryogenesis inCaenorhabditis elegans

Cited by 7 publications

References 0 publications

Analysis of smu-1, a Gene That Regulates the Alternative Splicing of unc-52 Pre-mRNA in Caenorhabditis elegans

Analysis of smu-1, a Gene That Regulates the Alternative Splicing of unc-52 Pre-mRNA in Caenorhabditis elegans

Proteomic approach to substrates related to MAPK pathway in 293T cells

Spermatogenetic Expression of RNA‐Binding Motif Protein 7, a Protein That Interacts With Splicing Factors

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