DOI: 10.1073/pnas.1635129100. Processive phosphorylation of alternative splicing factor/splicing factor 2

Processive phosphorylation of alternative splicing factor/splicing factor 2

10.1073/pnas.1635129100

Crossref journal-article

Proceedings of the National Academy of Sciences

Proceedings of the National Academy of Sciences (341)

Abstract

SR proteins, named for their multiple arginine/serine (RS) dipeptide repeats, are critical components of the spliceosome, influencing both constitutive and alternative splicing of pre-mRNA. SR protein function is regulated through phosphorylation of their RS domains by multiple kinases, including a family of evolutionarily conserved SR protein-specific kinases (SRPKs). The SRPK family of kinases is unique in that they are capable of phosphorylating repetitive RS domains with remarkable specificity and efficiency. Here, we carried out kinetic experiments specially developed to investigate how SRPK1 phosphorylates the model human SR protein, ASF/SF2. By using the start–trap strategy, we monitored the progress curve for ASF/SF2 phosphorylation in the absence and presence of an inhibitor peptide directed at the active site of SRPK1. ASF/SF2 modification is not altered when the inhibitor peptide (trap) is added with ATP (start). However, when the trap is added first and allowed to incubate for a specific delay time, the decrease in phosphate content of the enzyme–substrate complex follows a simple exponential decline corresponding to the release rate of SRPK1. These data demonstrate that SRPK1 phosphorylates a specific region within the RS domain of ASF/SF2 by using a fully processive catalytic mechanism, in which the splicing factor remains “locked” onto SRPK1 during RS domain modification.

Bibliography

Aubol, B. E., Chakrabarti, S., Ngo, J., Shaffer, J., Nolen, B., Fu, X.-D., Ghosh, G., & Adams, J. A. (2003). Processive phosphorylation of alternative splicing factor/splicing factor 2. Proceedings of the National Academy of Sciences, 100(22), 12601â12606.

Authors 8

Brandon E. Aubol (first)
Sutapa Chakrabarti (additional)
Jacky Ngo (additional)
Jennifer Shaffer (additional)
Brad Nolen (additional)
Xiang-Dong Fu (additional)
Gourisankar Ghosh (additional)
Joseph A. Adams (additional)

References 39 Referenced 95

Fu, X. D. (1995) RNA 1, 663–680.7585252 / RNA (1995)
10.1101/gad.6.5.837
10.1073/pnas.91.23.10824
10.1083/jcb.140.4.737
10.1139/o99-046
10.1073/pnas.181354098
10.1074/jbc.274.16.11125
10.1006/excr.1998.4083
10.1002/j.1460-2075.1996.tb00357.x
10.1038/369678a0
10.1093/emboj/17.21.6359
10.1101/gad.11.3.334
10.1093/nar/23.16.3260
10.1083/jcb.143.2.297
10.1016/S0960-9822(95)00060-1
10.1074/jbc.M100055200
10.1016/0076-6879(91)00127-I
10.1038/84178
10.1080/07391102.1983.10507477
10.1016/S0076-6879(80)64013-0
10.1006/jmbi.1993.1644
10.1111/j.1749-6632.1994.tb52803.x
10.1016/S0969-2126(01)00676-1
10.1016/0076-6879(95)62023-0
10.1073/pnas.89.22.10763
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10.1016/0003-2697(83)90660-7
Cao, W., Jamison, S. F. & Garcia-Blanco, M. A. (1997) RNA 3, 1456–1467.9404896 / RNA (1997)
10.1021/bi020233y
10.1016/S0955-0674(00)00212-X
10.1007/PL00000687
10.1242/jcs.113.11.1841
10.1017/S1355838200000960

Dates

Type	When
Created	21 years, 10 months ago (Oct. 29, 2003, 1:26 a.m.)
Deposited	3 years, 4 months ago (April 13, 2022, 2:12 a.m.)
Indexed	2 months ago (July 1, 2025, 9:46 p.m.)
Issued	21 years, 10 months ago (Oct. 10, 2003)
Published	21 years, 10 months ago (Oct. 10, 2003)
Published Online	21 years, 10 months ago (Oct. 10, 2003)
Published Print	21 years, 10 months ago (Oct. 28, 2003)

Funders 0

None

BibTeX

@article{Aubol_2003, title={Processive phosphorylation of alternative splicing factor/splicing factor 2}, volume={100}, ISSN={1091-6490}, url={http://dx.doi.org/10.1073/pnas.1635129100}, DOI={10.1073/pnas.1635129100}, number={22}, journal={Proceedings of the National Academy of Sciences}, publisher={Proceedings of the National Academy of Sciences}, author={Aubol, Brandon E. and Chakrabarti, Sutapa and Ngo, Jacky and Shaffer, Jennifer and Nolen, Brad and Fu, Xiang-Dong and Ghosh, Gourisankar and Adams, Joseph A.}, year={2003}, month=oct, pages={12601–12606} }

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