Worm Breeder's Gazette 10(2): 20

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.

Analysis of Phenotype and X-linked Gene Expression of sdc-2

Chad Nusbaum and Barbara Meyer

Mutations in the X-linked gene sdc-2 disrupt both the processes of 
sex determination and dosage compensation in XX animals (hence: sdc= 
sex determination and dosage compensation), but have no apparent 
effect on XO animals.  Mutant sdc-2 XX animals adopt the male modes of 
both sex determination and dosage compensation, resulting in 
masculinization and specific over-expression of X-linked transcripts, 
respectively.  Strong alleles exhibit XX specific lethality as 
homozygotes and in trans to deficiencies.
There are now twenty-two alleles of sdc-2.  They define an allelic 
series of increasing lethality and masculinization.  For strong 
alleles, XX animals are inviable.  Rare surviving XX homozygotes are 
Dpy and Tra.  For weak alleles, XX animals develop as Tra or 
intersexual animals or rare self fertile Egl hermaphrodites.  XX 
lethality and Tra phenotype vary with the strength of the allele.  A 
her-1 mutation can be used to block the Tra effect of an allele with 
substantial XX viability, producing a strain of her-1; 
phrodites.  Seven weak alleles allow some 
survival of XX animals and can be kept as homozygous strains in the 
presence of a her-1 mutation.  In order to look directly at dosage 
compensation defects of sdc-2 in XX animals, we looked at weaker 
alleles in her-1 homozygous strains.
We have done Northern blot analysis of RNA prepared from mixed-stage 
cultures of her-1(e1520); ) XX animals.  [Since XO 
her-1 animals are hermaphrodite, and since XO hermaphrodites in this 
strain may be healthier than XXs, 20 animals from the culture were 
verified genetically to be XX rather than XO.] An approximate twofold 
over-experession of X-linked mRNAs was seen.  This was observed for 
the X-linked trancript uvt-4 as normalized to the autosomal transcript 
act-1, and also with the X-linked transcript myo-2 as normalized to 
the autosomal trancript myo-1.sdc-2(y55) is an intermediate strength 
allele.  The majority of homozygous XX animals are masculinized and 
intersexual.  Egg counts of progeny from rare sdc-2(y55)/sdc-2(y55) XX 
homozygous hermaphrodites reveal that some lethality (about 5%) is 
observable, as either dead eggs or arrested L1 larvae.  6% of 
surviving adult animals are Egl hermaphrodites.  The remaining 94% are 
Tra and pseudomale, and many of these are very unhealthy.  They range 
from sterile intersexual animals with a vulval blip, through (often 
Dpy) animals of various intersexual character with deformed male tails,
all the way to animals that appear very close to wild-type males in 
phenotype.  The male structures of the latter can approach those of 
the wild-type male.  Animals have been observed under Nomarski optics 
with tail structures including 9 rays on each side, hook, spicules, 
and near wild type fan, and with wild type male gonad morphology.  
These animals can exhibit male mating behavior but fail to produce 
cross progeny.  Consistent with this, the gonads of these animals are 
seen to be packed with sperm, as if their mating defect resides in an 
inability to emit sperm.
Interestingly, a single mutant copy of her-1 is sufficient to block 
sdc-2(y55) masculinization.  Surviving sdc-2(y55)/sdc-2(y55) XX 
animals are 6% hermaphrodite, 94% Tra.  Adult her-1(e1520)/+; )
/sdc-2(y55) XX animals are 98% hermaphrodite, 2% Tra.  her-1(e1520); 
) XX animals are fully hermaphrodite, but are small,
dumpy, egg laying defective, and have protruding vulvae; phenotypes 
associated with mutations that disrupt hermaphrodite dosage 
compensation.  We have examined y55 in trans to nDf19 and to y39, a 
strong sdc-2 allele.  52% of y55/nDf19 animals survive and are Tra, 3% 
of these are self fertile.  y55/y39 animals are 15% viable and 
strongly Tra.
Three factor crosses involving dpy-6(e14) and lin-14(n179ts), and 
lin-14(n179ts) and unc-9(e101) now place sdc-2 within 0.1 map units to 
the right of lin-14.  In addition, two small deficiencies from the 
Ambros lab, maDf1 and maDf2, uncover both sdc-2 and lin-14 and fail to 
uncover egl-15, located 1 map unit to the left of lin-14 (M.  Stern 
personal communication) and sma-5, located 1.2 map units to the right. 
Both the strong allele y39 (XX lethal) and the intermediate strength 
allele y55 (XX Tra) have been mapped to within 0.2 map units of lin14. 
We have shown several alleles of both classes to be uncovered by the 
small deficiency maDf1.  This mapping helps to establish that 
mutations of both classes appear to occur at the same locus.  Further, 
as the lin-14 contig is now very large, this genetic position along 
with 4 gamma-radiation induced alleles and one allele generated in a 
mutator background, will facilitate cloning of sdc-2.