The Drosophila suppressor of sable protein binds to RNA and associates with a subset of polytene chromosome bands.
Mutations of the Drosophila melanogaster suppressor of sable [su(s)] gene, which encodes a 150-kDa nuclear protein [Su(s)], increase the accumulation of specific transcripts in a manner that is not well understood but that appears to involve pre-mRNA processing. Here, we report biochemical analysis of purified, recombinant Su(s) [rSu(s)] expressed in baculovirus and in Escherichia coli as maltose binding protein (MBP) fusions and immunocytochemical analysis of endogenous Su(s). This work has shown that purified, baculovirus-expressed rSu(s) binds to RNA in vitro with a high affinity and limited specificity. Systematic evolution of ligands by exponential enrichment was used to identify preferred RNA targets of rSu(s), and a large proportion of RNAs isolated contain a full or partial match to the consensus sequence UCAGUAGUCU, which was confirmed to be a high-affinity rSu(s) binding site. An MBP-Su(s) fusion protein containing the N-terminal third of Su(s) binds RNAs containing this sequence with a higher specificity than full-length, baculovirus-expressed rSu(s). The consensus sequence resembles both a cryptic 5' splice site and a sequence that is found near the 5' end of some Drosophila transcripts. Immunolocalization studies showed that endogenous Su(s) is distributed in a reticulated pattern in Drosophila embryo and salivary gland nuclei. In salivary gland cells, Su(s) is found both in the nucleoplasm and in association with a subset of polytene chromosome bands. Considering these and previous results, we propose two models to explain how su(s) mutations affect nuclear pre-mRNA processing.
Murray
M V
MV
Curriculum in Genetics and Molecular Biology, University of North Carolina, Chapel Hill 27599, USA.
Turnage
M A
MA
Williamson
K J
KJ
Steinhauer
W R
WR
Searles
L L
LL
D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D002875:Chromosomes / Q000378:metabolism; D003001:Cloning, Molecular; D004331:Drosophila melanogaster / Q000235:genetics* / Q000378:metabolism*; D017344:Genes, Insect; D007150:Immunohistochemistry; D019476:Insect Proteins / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D012313:RNA / Q000235:genetics / Q000378:metabolism*; D012322:RNA Precursors / Q000235:genetics / Q000378:metabolism; D012323:RNA Processing, Post-Transcriptional; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism
D019476:Insect Proteins; D009687:Nuclear Proteins; D012322:RNA Precursors; D011993:Recombinant Fusion Proteins; D012313:RNA