A coactivator of pre-mRNA splicing

Abstract

The nuclear matrix antigen recognized by the monoclonal antibody (mAb) B1C8 is a novel serine (S) and arginine (R)-rich protein associated with splicing complexes and is named here SRm160 (SR-related matrix protein of 160 kD). SRm160 contains multiple SR repeats, but unlike proteins of the SR family of splicing factors, lacks an RNA recognition motif. SRm160 and a related protein SRm300 (the 300-kD nuclear matrix antigen recognized by mAb B4A11) form a complex that is required for the splicing of specific pre-mRNAs. The SRm160/300 complex associates with splicing complexes and promotes splicing through interactions with SR family proteins. Binding of SRm160/300 to pre-mRNA is normally also dependent on U1 snRNP and is stabilized by U2 snRNP. Thus, SRm160/300 forms multiple interactions with components bound directly to important sites within pre-mRNA. The results suggest that a complex of the nuclear matrix proteins SRm160 and SRm300 functions as a coactivator of pre-mRNA splicing.[Key Words: Serine-arginine-rich splicing factor; small nuclear ribonucleoprotein particle; spliceosome; coactivator; nuclear matrix] Received September 30, 1997; revised version accepted January 29, 1998.Precursor mRNA (pre-mRNA) splicing occurs within the spliceosome, a ∼60S complex composed of four small nuclear ribonucleoprotein particles (U1, U2, U4/U6, U5 snRNPs) and multiple non-snRNP factors (for review, see Sharp 1994;Krä mer 1996). Although a wealth of biochemical and genetic data have provided detailed information on the role of specific spliceosomal components, it is not well understood how intron-exon segments are discriminated in vivo. To gain insights into this question, it will be important to determine how the splicing process is physically organized within the cell nucleus.In situ hybridization and immunocytochemistry studies have localized in the nucleus individual components involved in pre-mRNA synthesis and processing. The majority of splicing factors are both widely distributed throughout the nucleus, excluding nucleoli, and are also concentrated in two distinct types of foci, referred to as ''speckles'' and coiled bodies (for review, see Lamond and Carmo-Fonseca 1993;Lawrence et al. 1993;Spector 1993). Mammalian nuclei typically contain 20-50 speckles and 1-5 coiled bodies. Both of these structures concentrate all four snRNPs, but only speckles are enriched for the largest subunit of RNA polymerase II (Pol II LS; Bregman et al. 1995;Mortillaro et al. 1996), poly(A) + RNA (Carter et al. 1991;Huang et al. 1994), and essential non-snRNP splicing factors of the serine-arginine (SR)-family (Fu and Maniatis 1990;Spector et al. 1991). Fluorescence in situ hybridization studies have demonstrated that pre-mRNAs and spliced mRNAs of several specific cellular transcripts are located at the periphery or within speckles, indicating that a subset of pre-mRNAs are processed at these sites (Xing et al. , 1995. However, detection of nascent RNA synthesis by a brief pulse with (BrdU) or 3 H uridine reveals that the majority of na...

Cloning and characterization of two neural‐salient serine/arginine‐rich (NSSR) proteins involved in the regulation of alternative splicing in neurones

“…Recent studies have suggested that a coactivator(s) of pre-mRNA splicing in nuclear extracts is necessary for adequate splicing activity (Blencowe et al 1998).…”

The spatial targeting and nuclear matrix binding domains of SRm160

“…Two strong candidates for factors that might couple splicing components are Ser-Arg (SR)-related matrix protein of 160 kDa (SRm160) and SR-related matrix protein of 300 kDa (SRm300), two high molecular mass SR-related proteins (11)(12)(13)(14)(15). These proteins are bound more tightly to the nuclear matrix than other SR proteins, are binding partners, and are constituents of in vitro-assembled spliceosomes.…”

“…We suggest that SRm160 and SRm300 are key players in the recruitment of splicing factors to sites of splicing and in the assembly of presplicing complexes on the nuclear matrix, contributing to the greater efficiency of splicing in intact nuclei. Although both proteins participate in splicing reactions, SRm160 may be the more important for in vitro splicing, being required for the splicing of some RNA substrates (13,14). …”

Redundant enhancement of mouse constitutive androstane receptor transactivation by p160 coactivator family members

“…However, the nuclear speckles observed from SRC-1 and the round granules observed from SRC-2 and SRC-3 are morphologically similar to discrete subnuclear compartments that are associated with specific nuclear activities such as mRNA synthesis, pre-mRNA splicing [38,39,40]. Splicing factors such as SC-35 and SRm160 are primarily localized to nuclear foci referred to as splicing speckles or nuclear speckles [41,42]. Consistent with our observations, SRC-2/GRIP1 and SRC-3 have been found to associate with ND10 bodies, a small nuclear substructure containing several proteins that are implicated in the regulation of transcription, apoptosis, tumor suppression and the anti-viral response [43].…”