Blom7α Is a Novel Heterogeneous Nuclear Ribonucleoprotein K Homology Domain Protein Involved in Pre-mRNA Splicing That Interacts with SNEVPrp19-Pso4

Abstract

The removal of introns from pre-mRNA is performed by the spliceosome that stepwise assembles on the pre-mRNA before performing two catalytic steps. The spliceosome-associated CDC5L-SNEVPrp19-Pso4 complex is implicated in activation of the second catalytic step of pre-mRNA splicing, and one of its members, SNEVPrp19-Pso4, is also implicated in spliceosome assembly. To identify interaction partners of SNEVPrp19-Pso4, we have performed yeast two-hybrid screenings. Among the putative binding partners was a so far uncharacterized protein carrying two heterogeneous nuclear ribonucleoprotein K homology domains that we termed Blom7α. Blom7α is expressed in all tissues tested, and at least three splice variants exist. After confirming direct and physical interaction of SNEV and Blom7α, we investigated if it plays a functional role during pre-mRNA splicing. Indeed, Blom7α co-localizes and co-precipitates with splicing factors and pre-mRNA and is present in affinity-purified spliceosomes. More importantly, addition of Blom7α to HeLa nuclear extracts increased splicing activity in a dose-dependent manner. Furthermore, we tested if Blom7α influences splice site selection using two different minigene constructs. Indeed, both 5′- as well as 3′-site selection was altered upon Blom7α overexpression. Thus we suggest that Blom7α is a novel splicing factor of the K homology domain family that might be implicated in alternative splicing by helping to position the CDC5L-SNEVPrp19-Pso4 complex at the splice sites.

Inhibition of Pre-mRNA Splicing by a Synthetic Blom7α-Interacting Small RNA

“…As expected, full length Blom7α mainly localized to the nucleus of Hela cells, as we already described for the endogenous protein [6]. This was also true for all mutants containing the region between amino acids 480 and 549.…”

“…This was also true for all mutants containing the region between amino acids 480 and 549. However, mutants lacking this part and containing the KH motifs localized to nucleus, as well as cytoplasm, as was already described for Blom7γ, which is composed of the KH domains and the γ-specific C-terminus alone [6]. The cytoplasmic staining fully co-localized with RNA counterstained by Pyronin Y (Fig.…”

Mammalian splicing factor SF1 interacts with SURP domains of U2 snRNP-associated proteins

“…Moreover, KIAA0907 was found as a SF1 partner with very high confidence in all cell types tested. Roles in splicing for this protein, also termed BLOM7, have been described ( 60 ). It contains a KH domain, similar to a few other proteins found in the Y2H screens with a lower PBS.…”

“…Another protein with a high PBS found in all cell types analyzed is KIAA0907/BLOM7. It was initially identified in Y2H screens with a component of the human Prp19 complex and has been shown to increase splicing in HeLa cell nuclear extracts and regulate alternative splice site decisions ( 60 ). BLOM7 contains two KH domains, the second of which is comprised in the SF1 ΣSID.…”

BCAS2 is essential for Drosophila viability and functions in pre-mRNA splicing

“…In general, trans-acting factors that regulate alternative splicing include SR (Ser-Arg) proteins and heterogeneous nuclear ribonucleoproteins (hnRNPs) particles (Chen and Manley 2009). Recent studies showed that hPrp19 interacts directly with Blom7α, a novel hnRNP K homology domain-containing protein that can modulate the alternative 5 ′ and 3 ′ splice site choices of reporter minigenes (Grillari et al 2009). In addition, CDC5L/PLRG1 interacts with hnRNP M and affects the selection of alternative splice site (Lleres et al 2010).…”