DIFFERENTIAL PHOSPHORYLATIONS OF SPI-B AND SPI-1 TRANSCRIPTION FACTORS

Citation
C. Mao et al., DIFFERENTIAL PHOSPHORYLATIONS OF SPI-B AND SPI-1 TRANSCRIPTION FACTORS, Oncogene, 12(4), 1996, pp. 863-873
Citations number
55
Categorie Soggetti
Oncology,Biology,"Cell Biology
Journal title
ISSN journal
09509232
Volume
12
Issue
4
Year of publication
1996
Pages
863 - 873
Database
ISI
SICI code
0950-9232(1996)12:4<863:DPOSAS>2.0.ZU;2-H
Abstract
Spi-1/PU-1 and Spi-B are hematopoietic transcription factors which, in vitro, display similar affinities for DNA target sequences containing the consensus binding site 5'GGAA-3'. While the role of Spi-1 in the transcriptional regulation of B cell and myeloid specific genes has be en largely demonstrated, the biological function of Spi-B still remain s to be elucidated. Since Spi-B and Spi-1 are very divergent in their transactivator domain, these domains might acquire functional specific ity in vivo by interacting with different co-factors and/or by undergo ing different phosphorylations. First, we observed that casein kinase II phosphorylates Spi-B as well as Spi-1, in vitro, Then. by affinity chromatographies and in vitro kinase assays with fusion proteins betwe en glutathione-S-transferase and the transactivator domain of Spi-B, t wo kinases were identified on their ability to interact and phosphoryl ate this domain; the MAP kinase ERK1 and the stress activated protein kinase JNK1. The Threonine 56 was defined as the ERK1 phosphorylation site by using phosphoamino-acid analyses and a Spi-B mutant version wi th the substitution T56 to A56. Strikingly, ERK1 failed to phosphoryla te Spi-1, in vitro, whereas JNK1, like CK II, phosphorylated Spi-B and Spi-1. In addition, other purified Spi-B-kinase activities, unidentif ied as yet, display similar specificity than ERK1 for Spi-B versus Spi -1. Furthermore, the evident interaction of pRb protein with the trans activator domain of Spi-B in an unphosphorylated state disappeared whe n this domain was first phosphorylated in vitro either by ERK1 or by t he purified Spi-B-kinase activities. Our data revealed multiple phosph orylation sites within Spi-B whose some of them appeared specific for Spi-B versus Spi-1 and which may account for differential regulation o f their activities.