Effects of the U1C L13 mutation and temperature regulation of yeast commitment complex formation.
The U1 small nuclear ribonucleoprotein particle U1C protein has a zinc finger-like structure (C2H2 motif) at its N terminus, which is conserved from yeast to humans. Mutations of amino acid L13 within this domain rescue the essential function of the helicase protein Prp28p. Prp28p has been implicated in unwinding the 5' splice site (5'ss)-U1 small nuclear RNA (snRNA) base-pairing, to allow replacement of U1 snRNA with U6 snRNA during spliceosome assembly. The L13 phenotype has therefore been interpreted to indicate that WT U1C contributes to 5'ss-U1 snRNA stabilization by binding to the RNA duplex. We show here that an L13 mutant extract cannot form stable base-pairing at room temperature but is permissive for U1-5'ss base-pairing at low temperature. This phenotype is similar to that of a U1C-depleted extract, indicating that the U1C L13 mutation is a strong loss-of-function mutation. The two mutant extracts are unlike a WT extract, which undergoes stable pairing at room temperature but little or no pairing at low temperature. Taken together with previous results and the failure to observe a direct interaction of U1C with the U1-5'ss duplex, the data suggest that U1C contributes indirectly to stable U1-5'ss base-pairing under permissive conditions. A model is proposed to account for the L13 results.
Du
Hansen
H
Howard Hughes Medical Institute, Department of Biology, Brandeis University, MS008, 415 South Street, Waltham, MA 02454, USA.
Tardiff
Daniel F
DF
Moore
Melissa J
MJ
Rosbash
Michael
M
D007700:Kinetics; D012322:RNA Precursors / Q000235:genetics / Q000378:metabolism; D012331:RNA, Fungal / Q000235:genetics / Q000378:metabolism; D012342:RNA, Small Nuclear / Q000235:genetics / Q000378:metabolism; D017412:Ribonucleoprotein, U1 Small Nuclear / Q000235:genetics* / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics; D013816:Thermodynamics
D012322:RNA Precursors; D012331:RNA, Fungal; D012342:RNA, Small Nuclear; D017412:Ribonucleoprotein, U1 Small Nuclear; D029701:Saccharomyces cerevisiae Proteins; C405483:U1 small nuclear RNA