Worm Breeder's Gazette 14(3): 15 (June 1, 1996)

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.

Tissue-specific expression and RNA binding specificity of the C. elegans homologue of the spliceosome-associated protein SAP 49.

Yoshimi Tanaka, Atsunori Ohta, Hiroshi Sakamoto

Department of Biology, Faculty of Science, Kobe University, Rokkodai, Nadaku, Kobe 657, Japan

     We have isolated a C. elegans cDNA (accession no. U24189) that
encodes a putative 388 amino-acid RNA-binding protein with two RNA
recognition motifs (RRMs).  The RNA-binding protein shows extensive
similarity to the spliceosome-associated protein 49 (SAP 49) which has
been shown to be tightly involved in U2 snRNP function in mammalian cells
(Champion-Arnaud and Reed, Genes Dev. 8, 1974-83, 1994).  Surprisingly,
the cDNA also contains two additional ORFs upstream of the coding region
for the C. elegans SAP 49 homologue (cSAP49).  The product deduced from
the first ORF shows no homology with any known protein, whereas the
product of the second ORF encodes an actin-related protein.  Since very
short spacer regions containing the possible poly(A) and 3' splice sites
are present between these ORFs, and since these ORFs are continuous in the
genome sequence (CEC08B11; accession no. Z46676), it is suggested that
these ORFs are first transcribed in a single primary transcript and then
processed into three independent mRNAs by trans-splicing mechanism. 
Indeed, we confirmed three different mRNAs corresponding to the three
putative proteins by Northernblot analysis.
        By microinjection of the GFP- and LacZ-fusion constructs of cSAP49
under the control of the native promoter, we have found that it is
expressed at the restricted regions of the developing worm, mainly in the
nuclei of intestine tissues.  This expression pattern is puzzling because
the human SAP 49 is thought to bear the essential function in all cells. 
In addition, using an in vitro selection method, we have demonstrated that
cSAP49 possesses specific RNA binding ability which is attributed to the
second RRM.  Its RNA binding is in essentially a sequence-specific manner
but is affected by the surrounding sequence context, suggesting that
cSAP49 may recognize the secondary or tertiary structure as an additive
information.  We speculate that a possible target for the cSAP49 may be
the U2 snRNA since it is highly structured and contains a sequence similar
to a part of the consensus sequence of cSAP49-selected RNAs.  We are
currently examining whether cSAP49 really binds to the U2 snRNA in the
presence or absence of another U2-associated protein SAP 145, and whether
there would be other tissue-specific SAP 49 homologue in C. elegans.