Worm Breeder's Gazette 13(5): 66 (February 1, 1995)

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.


Kimberly C. Ferguson, Paul J. Heid, and Joel H. Rothman

Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 

      In screens for zygotic genes that act early in embryogenesis, we 
isolated two EMS-induced alleles (e2482 and w1) of a locus which we 
called zen-1 (1).   Both alleles cause cell migration defects beginning 
at the onset of gastrulation, a failure to undergo compaction and 
morphogenesis, and cell fate specification defects, suggesting an early 
zygotic requirement for this locus.
      By deficiency and polymorphism mapping we narrowed the region of
this  locus to ~400 kb within the unc-76 - dpy-21 interval.  Pools of
cosmids  in this region were scored for their ability to rescue embryonic 
lethality.  One cosmid, K07H12, was identified that rescued the 
lethality.  We found that K07H12 consists mostly of 1 kb repeats from  the
rrs-1 cluster.  rrs-1 consists of 110 tandemly repeated 1 kb units,  each
of which encodes both 5S rRNA and the trans-spliced leader RNA,  SL1. 
Surprisingly, a 1 kb repeat alone rescues the embryonic lethality  of both
of our alleles, suggesting that both mutations might be  deletions of the
rrs-1 cluster.  Indeed, Southern analysis of homozygous  embryos confirmed
that the cluster is completely deleted in both  mutants.
      To determine whether the embryonic lethality of these deletions
results  from the lack of SL1, 5S rRNA, or both, the genes encoding SL1
and 5S  rRNA were tested individually.  We found that SL1 is both
necessary and  sufficient for rescue of embryonic lethality; however,
hatching animals  generally arrest at early larval stages, presumably
owing to the lack of  zygotic 5S rRNA.  A 3 bp deletion in the 8 bp Sm
binding site of the SL1  outron [which is required for splicing in vitro
(2)] abolishes rescue,  suggesting that the ability of SL1 to be trans-
spliced is essential for  its rescuing activity.
      What is the fate of embryos deficient for SL1?  Mutations that
remove  SL1 in the rrs-1 cluster affect many processes during
embryogenesis.   Thus, SL1 may be essential for the activity of many RNAs. 
This is not  unexpected since SL1 is trans-spliced to ~56% of mRNAs in C.
elegans  (3).  However not all zygotic transcripts appear to require SL1,
as a  number of cell types are properly specified in these mutants (1).  
Perhaps some mRNAs which are normally trans-spliced to SL1 do not need 
SL1 to function properly, as has been demonstrated for rol-6 (4).     These
findings demonstrate that a trans-spliced leader, and probably the 
process of trans-splicing, are essential for early embryonic  development. 
Using our mutants we plan to address the normal role of  SL1 in metabolism
and/or expression of trans-spliced messages.  (For  example, is SL1
required for normal processing, transport, stability, or  translation of
trans-spliced RNAs?)

1) WBG 11: 4, p. 92; Worm meeting abstracts:  p. 145 (1991); p. 127 
2) Maroney, P.A., et. al. EMBO Journal 9, 3667-3673, (1990).
3) Spieth, J. et. al. Cell 73, 521-532 (1993).
4) Conrad, R. et. al. EMBO Journal 12, 1249-1255, (1993).