Worm Breeder's Gazette 14(1): 27 (October 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.

DPY-26 is a Novel Protein Associated with the X Chromosomes in XX Animals

Jason Lieb1, Beth Capowski2, Phil Meneely2,3, Barbara Meyer1

1 Department of Molecular and Cell Biology, University of California, Berkeley, California 94720
2 FHCRC, Seattle, WA 98104
3 Haverford College, Haverford, PA 19041

        C. elegans equalizes X-linked gene expression between XX
hermaphrodites and XO males by halving transcript levels from each of
the hermaphrodite X chromosomes. Although the mechanism of dosage
compensation is not known, molecular analysis of dpy-27 has yielded
significant insight. DPY-27 is a member of the evolutionarily conserved
SMC family of proteins, which are involved in chromosome condensation
and segregation. Moreover, DPY-27 is specifically localized to the X
chromosome in XX animals. The similarity of DPY-27 to SMC proteins and
its X localization suggest that dosage compensation is achieved through
changes in X chromosome structure. DPY-27, however, does not act alone.
sdc-1, sdc-2, sdc-3, dpy-21, dpy-26, dpy-27, dpy-28, and dpy-30 are also
required for the proper execution of dosage compensation.
        To further our understanding of dosage compensation we undertook
a molecular analysis of dpy-26. Homozygous mutant daughters of dpy-26
heterozygotes are dumpy and produce 4% male progeny. The XX progeny of
these homozygous mutants die as embryos or L1s, whereas XO animals
develop into normal males. The eight alleles of dpy-26 have
indistinguishable phenotypes. XX lethality is caused by the inability of
these animals to implement the hermaphrodite mode of dosage
compensation, leading to the overexpression of X-linked genes. Our
cloning and DNA sequence analysis has revealed that the predicted dpy-26
gene product is an acidic 1263 aa (142.5 kD) protein with no similarity
to any known motif, protein, or predicted protein sequence. The DNA
change in dpy-26(n199) is a C to T transition that results in a
premature termination of translation at codon 525. dpy-26 is
trans-spliced to both SL1 and SL2 leaders. Preliminary analysis suggests
that dpy-26 is the third gene in a three gene operon. Rescue of dpy-26
XX lethality is achieved with a subclone extending only 625 bp 5¹ of the
first codon, indicating that the upstream portion of the operon is not
required for rescue.
        Mouse polyclonal antibodies were raised against two DPY-26
polypeptides (aa 127-738 and aa 739-1263). Staining with either antibody
revealed that DPY-26 co-localizes precisely with DPY-27 in older (>50
cells) wild-type XX embryos, indicating that DPY-26 is associated with
the X chromosome. DPY-26 colocalizes with DPY-27 in all other embryos
studied as well. DPY-26 is not localized to the X chromosome in XO male
embryos or young (<50 cell) XX embryos, but is distributed diffusely
throughout the nucleus. This diffuse nuclear staining is also observed
in animals carrying either an sdc-2 or an sdc-3 mutation, revealing that
these genes are required for the proper localization of DPY-26 to X.
DPY-26 is mislocalized to the X chromosome of xol-1 XO embryos, which
have inappropriately activated the XX mode of dosage compensation.
DPY-26 X chromosome localization is unaffected by mutations in sdc-1 or
        dpy-26 and dpy-28 are necessary for the synthesis or stability
of DPY-27, since there is abundant dpy-27 mRNA but no DPY-27 protein in
dpy-26 or dpy-28 mutants. We find that DPY-27 and DPY-28 are probably
required for the stability of DPY-26, since there is no detectable
DPY-26 in either a dpy-27 or a dpy-28 mutant. Northern analysis to
determine dpy-26 transcript levels in these mutants is underway. These
data support the speculation that DPY-26, DPY-27, and DPY-28 are part of
a large dosage compensation protein complex associated specifically with
the X chromosome.
        There is one noteworthy difference between the distribution of
DPY-26 and DPY-27. In wild-type XX adults, DPY-27 is excluded from the
germline. As predicted by the Him phenotype of dpy-26 mutants, DPY-26 is
expressed in the germline. We will continue our study of dpy-26-induced
dosage compensation defects and meiotic non-disjunction in search of a
molecular explanation of these phenotypes.