Worm Breeder's Gazette 9(3): 57

These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.

Are Leader Sequences Spliced onto Actin mRNAs?

M. Krause and D. Hirsh

We have made gene-specific oligonucleotides to the 5' untranslated 
regions of actin mRNAs in order to map the transcriptional start sites 
of the four genes by primer extension. Primer extension dideoxy 
sequencing of actin gene 4 (X) mRNA confirmed earlier Sl-mapping 
results of a -41 to -43 transcriptional start site relative to the 
initiator AUG. The primer extension sequence was identical to the 
genomic sequence found upstream of actin gene 4. Primer extensions 
using actin gene 1 and/or 3 (V) and gene 2 (V) oligonucleotides 
yielded a unique 20 nucleotide sequence. This 20 nucleotide sequence 
is not the same as any of the genomic sequences adjacent to the 
primers used suggesting a splicing event during message maturation. 
This was not unexpected as each of these genespecific oligonucleotides 
abuts a consensus (genes 1 and 3) or near consensus (gene 2) TnCAG 
splice acceptor site. Actin genes 1 and 2 are oriented in opposite 
directions with their 5' ends toward each other. m e 20 nucleotide 
sequence which was derived from primer extensions is not in the 6 kb 
region containing actin genes 1 and 2.
In order to find the source of this unique 5' sequence on the actin 
mRNAs, an oligonucleotide homologous to the 20 nucleotide sequence was 
synthesized and used to probe Southern blots of genomic DNA digested 
with several enzymes. m e results suggest that the 20 nucleotide 
sequence is part of a 900 bp tandem repeat with a repeat copy number 
greater than 10. We have screened a C. MBL4 
library (provided by Chris Link) confirming the repetitive nature of 
the 20 nucleotide sequence and identifying phage of interest.
One possible interpretation of these results is that the 5' ends of 
actin 1,2 and 3 mRNAs arise by splicing leaders that are transcribed 
from a tandem repeat elsewhere in the genome. Precedents for splicing 
mRNA leader sequences derived from a separate RNA molecule exist in 
trypanosomes, mouse hepatitis virus and flu virus. Alternatively, the 
primer entension sequence from actin 1,2 and 3 mRNAs is a giant 
artifact. We are currently trying to decide between these