Conservation between yeast and man of a protein associated with U5 small nuclear ribonucleoprotein

Anderson, Gordon J.; Bach, Montserrat; Lührmann, Reinhard; Beggs, Jean D.

doi:10.1038/342819a0

Cited by 99 publications

(56 citation statements)

References 13 publications

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“…Cross-links of p220 and p70 to the branch region were associated with the B and C complexes. The p220 is almost certainly the U5-associated factor, the largest identified factor in the spliceosome and a homolog of the yeast PRP8 (Anderson et al 1989;GarciaBlanco et al 1990). In this and other studies, p220 has also been cross-linked to sites proximal to the 5' splice site (Wyatt et al 1992).…”

Section: Cross-linking To the Branch Sitementioning

confidence: 99%

Dynamic association of proteins with the pre-mRNA branch region.

et al. 1994

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The association of proteins with the branch site region during pre-mRNA splicing was probed using a novel methodology to site-specifically modify the pre-mRNA with the photo-reagent benzophenone. Three sets of proteins were distinguished by the kinetics of their associations with pre-mRNAs, by their association with discrete splicing complexes, and by their differing factor requirements. An early U1 snRNP-dependent cross-link of the branch region to a p80 species was followed by cross-links to p14, p35, and p150 polypeptides associated with the U2 snRNP-pre-mRNA complex. Concomitant with formation of the spliceosome, a rearrangement of protein factors about the branch region occurred, in which the p35 and p150 cross-links were replaced by p220 and p70 species. These results establish that the branch region is recognized in a dynamic fashion by multiple distinct proteins during the course of spliceosomal assembly.[Key Words: RNA splicing; photo-cross-linking; branch site; site-specific modifications] Received October 3, 1994; revised version accepted October 27, 1994.Pre-mRNA splicing occurs via two sequential transesterification reactions in a complex known as the spliceosome, a 60S entity that assembles on the pre-mRNA substrate in an ordered fashion through several stable complexes. The spliceosome includes the small nuclear ribonucleoprotein {snRNP)particles U1, U2, U4/6, and U5, as well as non-snRNP-associated splicing factors (for review, see Guthrie 1991;Lamm and Lamond 1993; Moore et al. 19931. Commitment of a pre-mRNA substrate to the splicing pathway involves the ATP-independent formation of the early or commitment complex, which contains tightly bound U1 snRNP as well as nonsnRNP protein factors, including U2AF bound near the branch region

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Section: Cross-linking To the Branch Sitementioning

confidence: 99%

Dynamic association of proteins with the pre-mRNA branch region.

et al. 1994

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“…Antibodies raised against the yeast PRP8 protein detect homologues in all species investigated, such as humans (13), mouse (P. Hodges and J. D. B., unpublished), Drosophila (14) and higher plants (15). Since the high molecular weight (>200 kDa) appears to be conserved among all homologues, the large size might be essential for the function of PRP8 protein.…”

Section: Introductionmentioning

confidence: 99%

Interaction of the yeast splicing factor PRP8 with substrate RNA during both steps of splicing

1995

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PRP8 protein of Saccharomyces cerevisiae interacts directly with pre-mRNA in spliceosomes, shown previously by UV-crosslinking. To analyse at which steps of splicing and with which precursor-derived RNA species the interaction(s) take place, UV-crosslinking was combined with PRP8-specific immunoprecipitation and the coprecipitated RNA species were analysed. Specific precipitation of intron-exon 2 and excised intron species was observed. PRP8 protein could be UV-crosslinked to pre-mRNA in PRP2-depleted spliceosomes stalled before initiation of the splicing reaction. Thus, the interaction of PRP8 protein with substrate RNA is established prior to the first transesterification reaction, is maintained during both steps of splicing and continues with the excised intron after completion of the splicing reaction. RNase Ti treatment of spliceosomes revealed that substrate RNA fragments of the 5' splice site region and the branchpoint-3' splice site region could be coimmunoprecipitated with PRP8 specific antibodies, indicating that these are potential sites of interaction for PRP8 protein with substrate RNA. Protection of the branchpoint-3' splice site region was detected only after step 1 of splicing. The results allow a first glimpse at the pattem of PRP8 protein-RNA interactions during splicing and provide a fundamental basis for future analysis of these interactions.

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“…In mammalian cells, about ten proteins have been found associated with the U4/U6 snRNP, none of which, however, is specific to this particle (13). In addition, several proteins have been shown to participate to the formation of the [U4/U6.U5] tri-snRNP, including the U5 snRNP specific protein, homologous to the yeast PRP8 protein (14)(15)(16)(17)(18)(19).…”

Section: Introductionmentioning

confidence: 99%