title: The unorthodox SNAP50 zinc finger domain contributes to cooperative promoter recognition by human SNAPC.
abstract: Human small nuclear RNA gene transcription by RNA polymerases II and III depends upon promoter recognition by the SNAPC general transcription factor. DNA binding by SNAPC involves direct DNA contacts by the SNAP190 subunit in cooperation with SNAP50 and SNAP43. The data presented herein shows that SNAP50 plays an important role in DNA binding by SNAPC through its zinc finger domain. The SNAP50 zinc finger domain contains 15 cysteine and histidine residues configured in two potential zinc coordination arrangements. Individual alanine substitution of each cysteine and histidine residue demonstrated that eight sites are important for DNA binding by SNAPC. However, metal binding studies revealed that SNAPC contains a single zinc atom indicating that only one coordination site functions as a zinc finger. Of the eight residues critical for DNA binding, four cysteine residues were also essential for both U1 and U6 transcription by RNA polymerase II and III, respectively. Surprisingly, the remaining four residues, although critical for U1 transcription could support partial U6 transcription. DNA binding studies showed that defects in DNA binding by SNAPC alone could be suppressed through cooperative DNA binding with another member of the RNA polymerase III general transcription machinery, TFIIIB. These results suggest that these eight cysteine and histidine residues perform different functions during DNA binding with those residues involved in zinc coordination likely performing a dominant role in domain stabilization and the others involved in DNA binding. These data further define the unorthodox SNAP50 zinc finger region as an evolutionarily conserved DNA binding domain.
authors.affiliation: Department of Microbiology & Molecular Genetics, Michigan State University, East Lansing, Michigan 48824, USA.
authors.forename: Gauri W
authors.initials: GW
authors.lastname: Jawdekar
authors.forename: Andrej
authors.initials: A
authors.lastname: Hanzlowsky
authors.forename: Stacy L
authors.initials: SL
authors.lastname: Hovde
authors.forename: Blanka
authors.initials: B
authors.lastname: Jelencic
authors.forename: Michael
authors.initials: M
authors.lastname: Feig
authors.forename: James H
authors.initials: JH
authors.lastname: Geiger
authors.forename: R William
authors.initials: RW
authors.lastname: Henry
mesh_terms: D000595:Amino Acid Sequence; D003545:Cysteine / Q000737:chemistry; D004268:DNA-Binding Proteins / Q000737:chemistry*; D006367:HeLa Cells; D006639:Histidine / Q000737:chemistry; D006801:Humans; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D017434:Protein Structure, Tertiary; D012313:RNA / Q000737:chemistry; D017386:Sequence Homology, Amino Acid; D014157:Transcription Factors / Q000737:chemistry* / Q000235:genetics*; D016335:Zinc Fingers
chemical_list: D004268:DNA-Binding Proteins; C106703:SNAPC protein complex, human; C093069:SNAPC1 protein, human; C101746:SNAPC3 protein, human; D014157:Transcription Factors; D006639:Histidine; D012313:RNA; D003545:Cysteine