Effect of PU.1 phosphorylation on interaction with NF-EM5 and transcriptional activation

Jagan M R Pongubala, Charles Van Beveren, Sujatha Nagulapalli, Michael Klemsz, Scott R. McKercher, Richard A. Maki, Michael L. Atchison

Research output: Contribution to journalArticle

225 Citations (Scopus)

Abstract

PU.1 recruits the binding of a second B cell-restricted nuclear factor, NF-EM5, to a DNA site in the immunoglobulin κ 3′ enhancer. DNA binding by NF-EM5 requires a protein-protein interaction with PU.1 and specific DNA contacts. Dephosphorylated PU.1 bound to DNA but did not interact with NF-EM5. Analysis of serine-to-alanine mutations in PU.1 indicated that serine 148 (Ser148) is required for protein-protein interaction. PU.1 produced in bacteria did not interact with NF-EM5. Phosphorylation of bacterially produced PU.1 by purified casein kinase II modified it to a form that interacted with NF-EM5 and that recruited NF-EM5 to bind to DNA. Phosphopeptide analysis of bacterially produced PU.1 suggested that Ser148 is phosphorylated by casein kinase II. This site is also phosphorylated in vivo. Expression of wild-type PU.1 increased expression of a reporter construct containing the PU.1 and NF-EM5 binding sites nearly sixfold, whereas the Ser148 mutant form only weakly activated transcription. These results demonstrate that phosphorylation of PU.1 at Ser148 is necessary for interaction with NF-EM5 and suggest that this phosphorylation can regulate transcriptional activity.

Original languageEnglish (US)
Pages (from-to)1622-1625
Number of pages4
JournalScience
Volume259
Issue number5101
StatePublished - Mar 12 1993
Externally publishedYes

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Serine
Transcriptional Activation
Phosphorylation
DNA
Casein Kinase II
Proteins
Phosphopeptides
Alanine
Immunoglobulins
B-Lymphocytes
Binding Sites
Bacteria
Mutation

ASJC Scopus subject areas

  • General

Cite this

Pongubala, J. M. R., Van Beveren, C., Nagulapalli, S., Klemsz, M., McKercher, S. R., Maki, R. A., & Atchison, M. L. (1993). Effect of PU.1 phosphorylation on interaction with NF-EM5 and transcriptional activation. Science, 259(5101), 1622-1625.

Effect of PU.1 phosphorylation on interaction with NF-EM5 and transcriptional activation. / Pongubala, Jagan M R; Van Beveren, Charles; Nagulapalli, Sujatha; Klemsz, Michael; McKercher, Scott R.; Maki, Richard A.; Atchison, Michael L.

In: Science, Vol. 259, No. 5101, 12.03.1993, p. 1622-1625.

Research output: Contribution to journalArticle

Pongubala, JMR, Van Beveren, C, Nagulapalli, S, Klemsz, M, McKercher, SR, Maki, RA & Atchison, ML 1993, 'Effect of PU.1 phosphorylation on interaction with NF-EM5 and transcriptional activation', Science, vol. 259, no. 5101, pp. 1622-1625.
Pongubala JMR, Van Beveren C, Nagulapalli S, Klemsz M, McKercher SR, Maki RA et al. Effect of PU.1 phosphorylation on interaction with NF-EM5 and transcriptional activation. Science. 1993 Mar 12;259(5101):1622-1625.
Pongubala, Jagan M R ; Van Beveren, Charles ; Nagulapalli, Sujatha ; Klemsz, Michael ; McKercher, Scott R. ; Maki, Richard A. ; Atchison, Michael L. / Effect of PU.1 phosphorylation on interaction with NF-EM5 and transcriptional activation. In: Science. 1993 ; Vol. 259, No. 5101. pp. 1622-1625.
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AB - PU.1 recruits the binding of a second B cell-restricted nuclear factor, NF-EM5, to a DNA site in the immunoglobulin κ 3′ enhancer. DNA binding by NF-EM5 requires a protein-protein interaction with PU.1 and specific DNA contacts. Dephosphorylated PU.1 bound to DNA but did not interact with NF-EM5. Analysis of serine-to-alanine mutations in PU.1 indicated that serine 148 (Ser148) is required for protein-protein interaction. PU.1 produced in bacteria did not interact with NF-EM5. Phosphorylation of bacterially produced PU.1 by purified casein kinase II modified it to a form that interacted with NF-EM5 and that recruited NF-EM5 to bind to DNA. Phosphopeptide analysis of bacterially produced PU.1 suggested that Ser148 is phosphorylated by casein kinase II. This site is also phosphorylated in vivo. Expression of wild-type PU.1 increased expression of a reporter construct containing the PU.1 and NF-EM5 binding sites nearly sixfold, whereas the Ser148 mutant form only weakly activated transcription. These results demonstrate that phosphorylation of PU.1 at Ser148 is necessary for interaction with NF-EM5 and suggest that this phosphorylation can regulate transcriptional activity.

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