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Differential intracellular localization of Sry proteins expressed in Rattus norvegicus
Author(s) -
Underwood Adam C,
Turner Monte E.,
Ely Daniel L.,
Milsted Amy
Publication year - 2009
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.23.1_supplement.1039.2
Subject(s) - testis determining factor , nuclear localization sequence , cytoplasm , microbiology and biotechnology , biology , nuclear protein , transfection , cell nucleus , western blot , intracellular , gene , subcellular localization , transcription factor , y chromosome , genetics
Sry is a transcription factor with a conserved HMG domain that facilitates DNA interactions and nuclear localization, through amino (n) and carboxy (c) terminal nuclear localization sequences (NLS). There are seven Sry loci on the SHR/Akr Y chromosome. Amino acid (aa) differences among Sry proteins are in the nNLS at aa 21 (His vs. Arg), cNLS at aa 76 (Pro vs. Thr), and in the presence/absence of 13 aa in a c‐ terminal activation motif. The objective of this study was to determine if these aa differences between proteins alter nuclear localization. Native, chimeric, mutated and truncated Sry sequences were transfected into CHO cells followed by immunocytochemistry and Western blot analysis of nuclear and cytoplasmic CHO cell fractions. Results showed that Sry2 exhibits both cytoplasmic and nuclear accumulation, while Sry1 and 3 localize only to the nucleus. Sry1 and 3 sequences containing the truncated or deleted activation region exhibit no reduction in nuclear localization, while mutations to the NLS result in cytoplasmic accumulation. Changing His to Arg at aa 21 of Sry1 or Sry3 results in nuclear localization patterns equivalent to native Sry2. Results demonstrate that aa variability in Sry proteins leads to different intracellular localization in vivo and may account for the different phenotypes observed in rats after delivery of exogenous Sry1, Sry2 or Sry3. Support from NIH HL71579 and the University of Akron.