scute (sis-b) Function in Drosophila Sex Determination

Deshpande, Girish; Stukey, Joseph; Schedl, Paul

doi:10.1128/mcb.15.8.4430

Cited by 50 publications

(35 citation statements)

References 56 publications

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“…3B, most of the Snf protein that could be immu- VOL. 16,1996 Sxl INTERACTS WITH SPLICING FACTORS 5039 noprecipitated by the Sxl antibody was present in fractions from the bottom half of the gradient. Since this Snf protein represents only a small subfraction of the total Snf protein, this outcome might help explain why the Snf antibody did not immunoprecipitate detectable amounts of the Sxl protein from total nuclear extracts.…”

Section: Resultsmentioning

confidence: 99%

“…This arginine residue is conserved in the mammalian U1A and U2BЉ pro- VOL. 16,1996 Sxl INTERACTS WITH SPLICING FACTORS 5043…”

Section: Resultsmentioning

confidence: 99%

“…In vitro studies have suggested a model in which Sxl regulates tra by competing with generic splicing factor VOL. 16,1996 Sxl INTERACTS WITH SPLICING FACTORS 5045 U2AF for binding to the poly(U) tract just upstream of the default (nonspecific) 3Ј splice site. As a consequence, U2 snRNP is not recruited to the default branch point but instead interacts with the downstream branch point of the femalespecific splice site.…”

Section: Discussionmentioning

confidence: 99%

“…Anti-Sxl monoclonal antibodies m114 and m5 are described by Bopp et al (10). Anti-Sis-b antibody 5A10 is described by Deshpande et al (16). Anti-Snf antibody 4G3 (23) was from the laboratory of Walther van Venrooij.…”

Section: Methodsmentioning

confidence: 99%

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Sex-lethal Interacts with Splicing Factors In Vitro and In Vivo

Deshpande

Samuels

Schedl

1996

Molecular and Cellular Biology

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The Drosophila sex determination gene Sex-lethal controls its own expression and the expression of downstream target genes such as transformer by regulating RNA splicing. Genetic and molecular studies have established that Sxl requires the product of another gene, snf, to autoregulate the splicing of its own transcripts. snf has recently been shown to encode a Drosophila U1 and U2 small nuclear ribonucleoprotein particle protein.In the work reported here, we demonstrate that the Sxl and Snf proteins can interact directly in vitro and that these two proteins are part of an RNase-sensitive complex in vivo which can be immunoprecipitated with the Sxl antibody. Unlike bulk Snf protein, which sediments slowly in sucrose gradients, the Snf protein associated with Sxl is in a large, rapidly sedimenting complex. Detailed characterization of the Sxl-Snf complexes from cross-linked extracts indicates that these complexes contain additional small nuclear ribonucleoprotein particle proteins and the U1 and U2 small nuclear RNAs. Finally, consistent with the RNase sensitivity of the Sxl-Snf complexes, Sxl transcripts can also be immunoprecipitated by Sxl antibodies. On the basis of the physical interactions between Sxl and Snf, we present a model for Sxl splicing regulation. This model helps explain how the Sxl protein is able to promote the sex-specific splicing of Sxl transcripts, utilizing target sequences that are distant from the regulated splice sites.In the fruit fly Drosophila melanogaster, the activity state of the binary switch gene, Sex-lethal (Sxl), is set early in development in response to the primary sex determination signal, the X chromosome-to-autosome ratio (15,17,20,32). Sxl is turned on in 2X/2A animals (females) by activating a posttranscriptional autoregulatory feedback loop (4,14). In this feedback loop, Sxl proteins promote their own synthesis by directing the female-specific splicing of Sxl transcripts originating from the Sxl maintenance promoter, Sxl-Pm (4). This autoregulatory feedback loop is responsible for maintaining the female-determined state during most of development. The Sxl autoregulatory feedback loop is not activated in 1X/2A animals (males), and Sxl-Pm transcripts are spliced in the nonproductive default mode. The critical difference between the Sxl mRNAs in the two sexes is the male-specific exon, L3 (Fig. 1). This exon contains in-frame translation stop signals that prematurely truncate an open reading frame which begins in exon L2 (5). Exon L3 is included in all male Sxl mRNAs, and these RNAs encode only very short, presumably nonfunctional polypeptides. In females, the Sxl protein mediates the skipping of the male exon, joining exon L2 to L4. This joining creates a long open frame which is predicted to encode protein species with sizes of about 35 kDa that contain two RNA recognition motif (RRM) domains (48,50).Sxl controls sexual differentiation by regulating gene cascades specifying different aspects of somatic sexual development (3, 28, 41). The best understood cascade is the tr...

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Section: Resultsmentioning

confidence: 99%

“…This arginine residue is conserved in the mammalian U1A and U2BЉ pro- VOL. 16,1996 Sxl INTERACTS WITH SPLICING FACTORS 5043…”

Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

See 2 more Smart Citations

Sex-lethal Interacts with Splicing Factors In Vitro and In Vivo

Deshpande

Samuels

Schedl

1996

Molecular and Cellular Biology

Self Cite

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“…, and the following transgenic alleles were used: P{w Antibodies: anti-Sxl m104 (1:10), anti-Sxl m114 (1:10), mouse anti-b-GAL (1:10) (Developmental Studies Hybridoma Bank, Iowa City, IA), anti-Snf 4G3 (1:10) (Flickinger and Salz 1994), anti-Scute5A10 (1:10) (Deshpande et al 1995), anti-U1-70K (1:6000) (Nagengast et al 2003), anti-U2AF50 (1:5000-50,000) (Rudner et al 1998), anti-U2AF38 (1:10,000) (Rudner et al 1996), anti-SKIPAb57 (1:2500) (Zhang et al 2003), anti-U5-40k (1:50,000) (Achsel et al 1998), anti-U5-116k (1:50,000) (Fabrizio et al 1997). Monoclonal line antiFl(2)d 9G2 was generated similarly to that described by Deshpande et al (1995) and used at a dilution of 1:10.…”

Section: De18mentioning

confidence: 99%

Functioning of the Drosophila Wilms'-Tumor-1-Associated Protein Homolog, Fl(2)d, in Sex-Lethal-Dependent Alternative Splicing

et al. 2008

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fl(2)d, the Drosophila homolog of Wilms'-tumor-1-associated protein (WTAP), regulates the alternative splicing of Sex-lethal (Sxl), transformer (tra), and Ultrabithorax (Ubx). Although WTAP has been found in functional human spliceosomes, exactly how it contributes to the splicing process remains unknown. Here we attempt to identify factors that interact genetically and physically with fl(2)d. We begin by analyzing the Sxl-Fl(2)d protein-protein interaction in detail and present evidence suggesting that the female-specific fl(2)d 1 allele is antimorphic with respect to the process of sex determination. Next we show that fl(2)d interacts genetically with early acting general splicing regulators and that Fl(2)d is present in immunoprecipitable complexes with Snf, U2AF50, U2AF38, and U1-70K. By contrast, we could not detect Fl(2)d complexes containing the U5 snRNP protein U5-40K or with a protein that associates with the activated B spliceosomal complex SKIP. Significantly, the genetic and molecular interactions observed for Sxl are quite similar to those detected for fl(2)d. Taken together, our findings suggest that Sxl and fl(2)d function to alter splice-site selection at an early step in spliceosome assembly. I N Drosophila melanogaster, sexual identity is initially determined by the X chromosome-to-autosome (A) ratio (Cline and Meyer 1996). The system that measures the X-to-A ratio turns on the Sex-lethal (Sxl) gene in female (XX) embryos by activating the Sxl establishment promoter, Sxl-Pe. In males (XY), this promoter is not activated and Sxl remains off. The establishment promoter is active for only a brief period in precellular blastoderm female embryos, and maintaining Sxl in the on state during the remainder of development in females depends upon an autoregulatory mechanism in which Sxl proteins direct their own synthesis by promoting the female-specific splicing of Sxl pre-mRNAs transcribed from the Sxl maintenance promoter (SxlPm). In this autoregulatory loop, Sxl proteins bind to multiple sites in the large introns located upstream and downstream of the male-specific third exon and promote the joining of the 59 splice site of exon 2 to the 39 splice site of exon 4, skipping exon 3. The translation of the resulting Sxl mRNA ensures the maintenance of female identity by providing a continuous source of Sxl protein. In males, the third exon is incorporated into the Sxl mRNA by the default splicing machinery. The male-specific exon has several in-frame stop codons that prematurely truncate the Sxl open reading frame, which begins in exon 2, resulting in the production of a truncated, nonfunctional polypeptide. Sxl controls the majority of female development by regulating the female-specific splicing of transformer (tra) pre-mRNA. This female-specific Tra protein (Tra F ) then regulates the alternative splicing of the transcription factors doublesex and fruitless. In addition, Sxl expression is required for female viability because it turns off the gene msl-2. MSL-2 is a component of the male-speci...

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The role of class I HLH genes in neural development—have they been overlooked?

Heng

Tan

2003

BioEssays

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Helix-loop-helix (HLH) genes encode for transcription factors affecting a whole variety of developmental programs, including neurogenesis. At least seven functional classes (denoted I to VII) of HLH genes exist, (1) with subclass members exhibiting homo- and heterodimerisation for proper DNA binding and transcriptional regulation of downstream target genes. In the developing nervous system, members of class II, V and VI have been most extensively studied concerning their roles in neural programming. In contrast, the function of class I proteins (such as E12 and E47) is poorly defined and the orthodox view relegates them to general dimerisation duties that are necessary for the activity of the other classes. However, closer scrutiny of the spatiotemporal expression patterns of class I factors, combined with recent biochemical evidence, would suggest that class I proteins possess specific functions during early neural differentiation. This essay supports this possibility, in addition to putting forward the hypothesis that, outside their general dimerisation activity, class I genes have independent roles in regulating neurogenesis.

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scute (sis-b) Function in Drosophila Sex Determination

Cited by 50 publications

References 56 publications

Sex-lethal Interacts with Splicing Factors In Vitro and In Vivo

Sex-lethal Interacts with Splicing Factors In Vitro and In Vivo

Functioning of the Drosophila Wilms'-Tumor-1-Associated Protein Homolog, Fl(2)d, in Sex-Lethal-Dependent Alternative Splicing

The role of class I HLH genes in neural development—have they been overlooked?

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