Purification and cDNA cloning of HeLa cell p54^nrb, a nuclear protein with two RNA recognition motifs and extensive homology to human splicing factor PSF andDrosophilaNONA/BJ6

Abstract

While searching for a human homolog of the S.cerevisiae splicing factor PRP18, we found a polypeptide that reacted strongly with antibodies against PRP18. We purified this polypeptide from HeLa cells using a Western blot assay, and named it p54nrb (for nuclear RNA-binding protein, 54 kDa). cDNAs encoding p54nrb were cloned with probes derived from partial sequence of the purified protein. These cDNAs have identical coding sequences but differ as a result of alternative splicing in the 5' untranslated region. The cDNAs encode a 471 aa polypeptide that contains two RNA recognition motifs (RRMs). Human p54nrb has no homology to yeast PRP18, except for a common epitope, but is instead 71% identical to human splicing factor PSF within a 320 aa region that includes both RRMs. In addition, both p54nrb and PSF are rich in Pro and Gln residues outside the main homology region. The Drosophila puff-specific protein BJ6, one of three products encoded by the alternatively spliced no-on-transient A gene (nonA), which is required for normal vision and courtship song, is 42% identical to p54nrb in the same 320 aa region. The striking homology between p54nrb, PSF, and NONA/BJ6 defines a novel phylogenetically conserved protein segment, termed DBHS domain (for Drosophila behavior, human splicing), which may be involved in regulating diverse pathways at the level of pre-mRNA splicing.

An actin–ribonucleoprotein interaction is involved in transcription by RNA polymerase II

“…10 and references therein), and the conformational state of the actin involved in transcription remains to be investigated. Hrp65 belongs to a family of evolutionarily conserved proteins characterized by the presence of a DBHS domain (27). The DBHS group includes the mammalian proteins PSF, p54 nrb ͞ NonO, and PSP1 and the Drosophila protein NonA͞Bj6 (reviewed in ref.…”

Fusion of splicing factor genes PSF and NonO (p54nrb) to the TFE3 gene in papillary renal cell carcinoma

“…For both 5' and 3'-RACE, sequences were obtained that diverged from known TFE3 sequences at the exon 3-exon 4 junction (eg, the 5'-RACE product is shown in Figure 2b). Database searches of the new sequences present in the 5'-RACE and 3'-RACE products revealed that they exactly matched adjacent regions of a gene designated NonO (p54 nrb ) that is closely related to PSF (Dong et al, 1993;Yang et al, 1993). The 471 amino acid NonO (p54 nrb ) protein contains several distinct domains ( Figure 3) including (i) a short N-terminal sequence composed entirely of histidine, glutamine and proline residues, (ii) a helix ± turn ± helix domain¯anked by charged amino acids that is responsible for binding to the octamer sequence in double stranded DNA and (iii) a short C-terminal proline-rich region.…”

“…The 471 amino acid NonO (p54 nrb ) protein contains several distinct domains ( Figure 3) including (i) a short N-terminal sequence composed entirely of histidine, glutamine and proline residues, (ii) a helix ± turn ± helix domain¯anked by charged amino acids that is responsible for binding to the octamer sequence in double stranded DNA and (iii) a short C-terminal proline-rich region. The NonO (p54 nrb ) protein has a region of 320 amino acids with 71% identity and 7% similarity to a 320aa region of PSF (Patton et al, 1993;Dong et al, 1993;Yang et al, 1993). This same region of NonO (p54 nrb ) exhibits 42% identity and 7% similarity to a 321aa region of the 700aa Drosophila NonA diss protein.…”

The Splicing Factor PSF is Part of a Large Complex That Assembles in the Absence of pre-mRNA and Contains All 5 snRNPs

“…14,[16][17][18][19][20][21][22][23] It is comprised of an N-terminal glycine-rich domain, a proline/glutamine (P/Q) rich domain, two RNA recognition motifs (RRMs), and a C-terminal region with two nuclear localization signals. A closely related protein with a similar domain structure, p54 nrb (and its mouse homolog nonO) 24,25 has been found to associate with PSF. 23,[26][27][28] Together, these proteins interact with the C-terminal domain (CTD) of RNA Pol II 29 and contact 5' splice sites within large transcription/splicing complexes, 30 consistent with coupling between transcription and splicing.…”