The snRNP-free U1A (SF-A) complex(es): Identification of the largest subunit as PSF, the polypyrimidine-tract binding protein-associated splicing factor

Lutz, Carol S.; Cooke, Charles L.; O’Connor, J. P.; Kobayashi, Ryûji; Alwine, James C.

doi:10.1017/s1355838298981183

Cited by 40 publications

(43 citation statements)

References 23 publications

(15 reference statements)

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“…In the de-repression experiments, U1A is present in the context of other endogenous proteins in the various cell extracts, and the inhibitory effect could be mediated or modulated by accessory proteins. Others have reported that RNA-free U1A is present in a complex with other proteins (O'Connor et al 1997;Lutz et al 1998). We therefore considered the possibility that U1A that binds mRNA might also be in complex with other proteins that might modulate its activity on that mRNA.…”

Section: Discussionmentioning

confidence: 99%

“…A decrease in the ratio of U1A to the snRNP associated protein, B¢ has been observed during differentiation, suggesting that less U1A is available in differentiated cells (Milcarek et al 2003). Furthermore, U1A has been reported to exist in a complex with other proteins, which may modulate its effect (O'Connor et al 1997;Lutz et al 1998). Therefore, a thorough investigation of the availability of U1A to inhibit poly(A) addition during B cell differentiation is necessary to understand the mechanism for the decrease in the inhibitory effect of U1A during differentiation.…”

Section: Introductionmentioning

confidence: 99%

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Non-snRNP U1A levels decrease during mammalian B-cell differentiation and release the IgM secretory poly(A) site from repression

Gunderson²,

Phillips

2005

RNA

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A regulated shift from the production of membrane to secretory forms of Immunoglobulin M (IgM) mRNA occurs during B cell differentiation due to the activation of an upstream secretory poly(A) site. U1A plays a key role in inhibiting the expression of the secretory poly(A) site by inhibiting both cleavage at the poly(A) site and subsequent poly(A) tail addition. However, how the inhibitory effect of U1A is alleviated in differentiated cells, which express the secretory poly(A) site, is not known. Using B cell lines representing different stages of B cell differentiation, we show that the amount of U1A available to inhibit the secretory poly(A) site is reduced in differentiated cells. Undifferentiated B cells have more total U1A than differentiated cells and a greater proportion of this is not associated with the U1snRNP. We show that this is available to inhibit poly(A) addition at the secretory poly(A) site using cold competitor RNA oligos to de-repress poly(A) addition in nuclear extracts from the respective cell lines. In addition, endogenous non-snRNP associated U1A-immunopurified from the different cell lines-inhibits poly(A) polymerase activity proportional to U1A recovered, suggesting that available U1A level alone is responsible for changes in its inhibitory effect at the secretory IgM poly (A) site.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Non-snRNP U1A levels decrease during mammalian B-cell differentiation and release the IgM secretory poly(A) site from repression

Gunderson²,

Phillips

2005

RNA

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“…Interestingly, we also identified SNRPA (U1A), a bifunctional protein primarily involved in pre-mRNA splicing but also shown to function in regulated polyadenylation. We additionally identified NONO (p54 nrb ), SFPQ (PSF), and PTBP1 (PTB), three factors known to bind snRNP-free U1A and function with it in polyadenylation (48,52,53). In addition to factors involved in polyadenylation, we identified Gemin5, the RNA-binding protein of the SMN complex (54).…”

Section: Identification Of Proteins That Bind the Polyadenylation Sitmentioning

confidence: 99%

U1A Regulates 3′ Processing of the Survival Motor Neuron mRNA

Workman

Veith

Battle

2014

Journal of Biological Chemistry

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Background: Insufficient expression of survival motor neuron (SMN), a protein involved in small nuclear ribonucleoprotein (snRNP) biogenesis, causes spinal muscular atrophy (SMA). Results: The U1A protein binds to and inhibits polyadenylation and cleavage of the SMN pre-mRNA. Conclusion: The U1A protein is a critical regulator of SMN expression. Significance: Understanding of the regulation of SMN expression is fundamental for developing treatments for SMA.

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“…PSF was first identified as a factor that associates with polypyrimidine tract-binding protein and was shown to be required at an early step in spliceosome assembly (61). PSF was also identified as a component of a small nuclear RNP-free complex (SF-A) that contains U1 small nuclear RNP-specific protein A and is implicated in 3Ј-end processing (27). PSF and p54nrb share considerable sequence homology and have been shown to bind the C-terminal domain of the largest subunit of RNA polymerase II (32).…”

Section: Cf I M 68 Interacts With the 25-kda Protein Via Its Rrm-mentioning

confidence: 99%

Distinct Sequence Motifs within the 68-kDa Subunit of Cleavage Factor Im Mediate RNA Binding, Protein-Protein Interactions, and Subcellular Localization

Dettwiler

Aringhieri

Cardinale

et al. 2004

Journal of Biological Chemistry

138

219

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Eukaryotic mRNA precursors (pre-mRNAs) 1 are synthesized and processed in the nucleus prior to their export to the cytoplasm where they serve as templates for protein synthesis. Transcription is coupled spatially and temporally to the capping of the pre-mRNA at the 5Ј-end, splicing, and 3Ј-end formation. The mature 3Ј-ends of most eukaryotic mRNAs are generated by endonucleolytic cleavage of the primary transcript followed by the addition of a poly(A) tail to the upstream cleavage product (for reviews see Refs. 1 and 2). In mammals, these reactions are catalyzed by a large multicomponent complex that is assembled in a cooperative manner on specific cis-acting sequence elements in the pre-mRNA. The cleavage and polyadenylation specificity factor (CPSF) (3) recognizes the highly conserved hexanucleotide AAUAAA, whereas the cleavage stimulation factor (CstF) (4) binds a more degenerate GUor U-rich element downstream of the poly(A) site. It has been suggested that in vivo CPSF and CstF may become associated with each other prior to pre-mRNA binding, recognizing the two elements in a concerted manner (5). In addition, the cleavage reaction requires mammalian cleavage factor I (CF I m ), cleavage factor II m (CF II m ), and poly(A) polymerase (PAP). After the first step of 3Ј-end processing, CPSF remains bound to the upstream cleavage fragment and tethers PAP to the 3Ј-end of the pre-mRNA (6). In the presence of the nuclear poly(A)-binding protein 1 (PABPN1), PAP elongates the poly(A) tail in a processive manner (6). These factors are both necessary and sufficient to reconstitute cleavage and polyadenylation in vitro. However, the other proteins involved in either transcription, such as the C-terminal domain of RNA polymerase II, or capping (nuclear cap-binding complex) and splicing (U2AF65) have been shown to greatly enhance the efficiency of the first step of the reaction (7-9).Three major polypeptides of 25, 59, and 68 kDa and one minor polypeptide of 72 kDa copurify with CF I m activity from HeLa cell nuclear extract (10). Reconstitution of CF I m activity with recombinant proteins suggests that CF I m is a heterodimer consisting of the 25-kDa subunit and one of the larger polypeptides (11). All of the three larger proteins appear to be highly related in their amino acid sequence. Moreover, all of the CF I m subunits are only present in metazoan organisms. The primary sequence of the 25-kDa polypeptide contains a NUDIX motif (12). The amino acid composition of the 68-kDa protein has a domain organization that is reminiscent of spliceosomal SR proteins. Members of the SR family of splicing factors contain one or more N-terminal RNA recognition motifs (RRMs) that function in sequence-specific RNA binding and a C-terminal domain rich in alternating arginine and serine residues, referred to as RS domain, which is required for proteinprotein interactions with other RS domains (13). In the 68-kDa protein, the RRM and the RS-like domain are separated by a

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The snRNP-free U1A (SF-A) complex(es): Identification of the largest subunit as PSF, the polypyrimidine-tract binding protein-associated splicing factor

Cited by 40 publications

References 23 publications

Non-snRNP U1A levels decrease during mammalian B-cell differentiation and release the IgM secretory poly(A) site from repression

Non-snRNP U1A levels decrease during mammalian B-cell differentiation and release the IgM secretory poly(A) site from repression

U1A Regulates 3′ Processing of the Survival Motor Neuron mRNA

Distinct Sequence Motifs within the 68-kDa Subunit of Cleavage Factor Im Mediate RNA Binding, Protein-Protein Interactions, and Subcellular Localization

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