Structural data suggest that human SNRP27 M141 and SNU-66 H734have a stacking interaction at the base of the U6 snRNA ACAGAGA box in
the pre-B complex. Previously, we found that mutations in C. elegans at
SNRP-27 M141 promote changes in cryptic 5’ splice site choice and
alternative 5’ss usage at native alternative splicing events 1
. We wanted to
better understand if the potential interaction between SNRP-27 M141 and
SNU-66 H765 (the C. elegans equivalent position to human SNU66 H734)
contributes to maintaining the 5’ splice site identity during spliceosome
assembly. Mutations at SNU-66 H765 change alternative 5’ splice sites usage
genome wide. SNU-66(H765G) is a suppressor of unc-73(e936) 5’ splice site
cryptic splicing, similar to SNRP-27 (M141T). Many of the alternative 5’
splicing events affected by SNU-66 (H765G) and (H765L) overlapped with
those affected SNRP-27 (M141T). Double mutants of snrp-27(M141T) with
snu-66(H765G) are homozygous lethal. Our findings show a new role for
SNU-66 in 5’ splice site selection in a similar way as SNRP-27 M141T.
Mutations at SNRP-27 M141 and SNU-66 H765 allow the spliceosome to
choose alternative 5’ splice sites with weaker consensus in the proximity of a
stronger consensus 5’ss. While we observed alternative 5’ splicing events that
could be promoted either upstream or downstream of the stronger 5’ss in these
mutants, the mechanism by which SNRP-27 M141 and SNU-66 H765
DocuSign Envelope ID: 13C88DBB-B62D-4F32-A3B3-56B16F7842AC
mutants affect 5’ splice sites is dependent on both the presence of a weaker
consensus 5’ss in close proximity and other factors (perhaps splicing factor
binding sites nearby) that are important for determining the directionality of
alternative splicing.