Sex ratio adjustment in relation to paternal attractiveness in a wild bird population.

When the relative fitness of sons and daughters differs, sex-allocation theory predicts that it would be adaptive for individuals to adjust their investment in different sexes of offspring. Sex ratio adjustment by females in response to the sexual attractiveness of their mate would be an example of this. In vertebrates the existence of this form of sex ratio adjustment is controversial and may be confounded with sex-biased mortality, particularly in sexually size-dimorphic species. Here we use PCR amplification of a conserved W-chromosome-linked gene to show that the sex ratio within broods of a natural population of sexually size-monomorphic collared flycatchers Ficedula albicollis is related to the size of their father's forehead patch, a heritable secondary sexual character implicated in female choice. Experimental manipulations of paternal investment, which influence the size of his character in future breeding attempts, result in corresponding changes in the sex ratio of offspring born to males in those breeding attempts. In contrast, manipulations of maternal investment have no effect on future sex ratios, and there is no relationship between variables predicting the reproductive value of the brood and nestling sex ratio. Analysis of recruitment of offspring reveals similar patterns of sex ratio bias. The results suggest that female collared flycatchers be able to adjust the sex ratio of eggs ovulated in response to the phenotype of their mate. This finding is most consistent with "genetic quality" models of sexual selection.

Loss of sequences 3' to the testis-determining gene, SRY, including the Y pseudoautosomal boundary associated with partial testicular determination.

The condition termed 46,XY complete gonadal dysgenesis is characterized by a completely female phenotype and streak gonads. In contrast, subjects with 46,XY partial gonadal dysgenesis and those with embryonic testicular regression sequence usually present ambiguous genitalia and a mix of Müllerian and Wolffian structures. In 46,XY partial gonadal dysgenesis gonadal histology shows evidence of incomplete testis determination. In 46,XY embryonic testicular regression sequence there is lack of gonadal tissue on both sides. Various lines of evidence suggest that embryonic testicular regression sequence is a variant form of 46,XY gonadal dysgenesis. The sex-determining region Y chromosome gene (SRY) encodes sequences for the testis-determining factor. To date germ-line mutations in SRY have been reported in approximately 20% of subjects with 46,XY complete gonadal dysgenesis. However, no germ-line mutations of SRY have been reported in subjects with the partial forms. We studied 20 subjects who presented either 46,XY partial gonadal dysgenesis or 46,XY embryonic testicular regression sequence. We examined the SRY gene and the minimum region of Y-specific DNA known to confer a male phenotype. The SRY-open reading frame (ORF) was normal in all subjects. However a de novo interstitial deletion 3' to the SRY-ORF was found in one subject. Although it is possible that the deletion was unrelated to the subject's phenotype, we propose that the deletion was responsible for the abnormal gonadal development by diminishing expression of SRY. We suggest that the deletion resulted either in the loss of sequences necessary for normal SRY expression or in a position effect that altered SRY expression. This case provides further evidence that deletions of the Y chromosome outside the SRY-ORF can result in either complete or incomplete sex reversal.

Patterns of kinship in groups of free-living sperm whales (Physeter macrocephalus) revealed by multiple molecular genetic analyses.

Mature female sperm whales (Physeter macrocephalus) live in socially cohesive groups of 10-30, which include immature animals of both sexes, and within which there is communal care of the young. We examined kinship in such groups using analyses of microsatellite DNA, mitochondrial DNA sequence, and sex-linked markers on samples of sloughed skin collected noninvasively from animals in three groups off the coast of Ecuador. Social groups were defined through photographic identification of individuals. Each group contained about 26 members, mostly female (79%). Relatedness was greater within groups, as compared to between groups. Particular mitochondrial haplotypes were characteristic of groups, but all groups contained more than one haplotype. The data are generally consistent with each group being comprised of several matrillines from which males disperse at about the age of 6 years. There are indications of paternal relatedness among grouped individuals with different mitochondrial haplotypes, suggesting long-term associations between different matrilines.

Multiplex genotype determination at a large number of gene loci.

To facilitate large-scale genotype analysis, an efficient PCR-based multiplex approach has been developed. For simultaneously amplifying the target sequences at a large number of genetic loci, locus-specific primers containing 5' universal tails are used. Attaching the universal tails to the target sequences in the initial PCR steps allows replacement of all specific primers with a pair of primers identical to the universal tails and converts the multiplex amplification into "uniplex." Simultaneous amplification of 26 genetic loci with this approach is described. The multiplex amplification can be coupled with genotype determination. By incorporating a single-base mismatch between a primer and the template into the target sequences, a polymorphic site can be converted into a desirable restriction fragment length polymorphism when it is necessary. In this way, the allelic PCR products for the polymorphic loci can be discriminated by gel electrophoresis after restriction enzyme digestion. In this study, 32 loci were typed in such a multiplex way.

Lens complementation system for the genetic analysis of growth, differentiation, and apoptosis in vivo.

A genetic approach has been established that combines the advantages of blastocyst complementation with the experimental attributes of the developing lens for the functional analysis of genes governing cellular proliferation, terminal differentiation, and apoptosis. This lens complementation system (LCS) makes use of a mutant mouse strain, aphakia (ak), homozygotes of which fail to develop an ocular lens. We demonstrate that microinjection of wild-type embryonic stem (ES) cells into ak/ak blastocysts produces chimeras with normal ES-cell-derived lenses and that microinjection of Rb-/- ES cells generates an aberrant lens phenotype identical to that obtained through conventional gene targeting methodology. Our determination that a cell autonomous defect underlies the aphakia condition assures that lenses generated through LCS are necessarily ES-cell-derived. LCS provides for the rapid phenotypic analysis of loss-of-function mutations, circumvents the need for germ-line transmission of null alleles, and, most significantly, facilitates the study of essential genes whose inactivation is associated with early lethal phenotypes.

Molecular markers reveal cryptic sex in the human pathogen Coccidioides immitis.

Coccidioides immitis, cause of a recent epidemic of "Valley fever" in California, is typical of many eukaryotic microbes in that mating and meiosis have yet to be reported, but it is not clear whether sex is truly absent or just cryptic. To find out, we have undertaken a population genetic study using PCR amplification, screening for single-strand conformation polymorphisms, and direct DNA sequencing to find molecular markers with nucleotide-level resolution. Both population genetic and phylogenetic analyses indicate that C. immitis is almost completely recombining. To our knowledge, this study is the first to find molecular evidence for recombination in a fungus for which no sexual stage has yet been described. These results motivate a directed search for mating and meiosis and illustrate the utility of single-strand conformation polymorphism and sequencing with arbitrary primer pairs in molecular population genetics.

Localization of the mouse gene releasing sex-limited expression of Slp.

To probe genetic variation in the regulation of sexual dimorphism, we have characterized the mouse protein Slp, coded by the gene sex-limited protein (Slp). Slp expression in many strains is limited to males and is androgen-dependent. However, female expression is also observed in rare strains, due to nonlinked gene(s) termed regulator of sex-limitation (rsl). In this report we demonstrate that female expression of Slp results from homozygous recessive allele(s) at a single autosomal locus that maps to a 2.2-centimorgan interval on chromosome 13. This conclusion was supported by extensive genetic analyses including the use of polymorphic microsatellites to type numerous backcross progeny and a recombinant inbred series and to identify the congenic interval in three independently derived congenic strains. Four attractive candidate genes were identified by the localization of rsl. Interestingly, rsl was found not only to enable expression in females but to also increase expression in males. The findings suggest that the expression of Slp and perhaps other sexually dimorphic proteins is regulated by two pathways, one that is dependent upon rsl but not androgens and another that is rsl-independent but requires androgens.

Isolation and characterization of a pseudoautosomal region-specific genetic marker in C57BL/6 mice using genomic representational difference analysis.

Representational difference analysis was used to identify strain-specific differences in the pseudoautosomal region (PAR) of mouse X and Y chromosomes. One second generation (C57BL/6 x Mus spretus) x Mus spretus interspecific backcross male carrying the C57BL/6 (B6) PAR was used for tester DNA. DNA from five backcross males from the same generation that were M. spretus-type for the PAR was pooled for the driver. A cloned probe designated B6-38 was recovered that is B6-specific in Southern analysis. Analysis of genomic DNA from several inbred strains of laboratory mice and diverse Mus species and subspecies identified a characteristic Pst I pattern of fragment sizes that is present only in the C57BL family of strains. Hybridization was observed with sequences in DBA/2J and to a limited extent with Mus musculus (PWK strain) and Mus castaneus DNA. No hybridization was observed in DNA of different Mus species, M. spretus, M. hortulanus, and M. caroli. Genetic analyses of B6-38 was conducted using C57BL congenic males that carry M. spretus alleles for distal X chromosome loci and the PAR and outcrosses of heterozygous congenic females with M. spretus. These analyses demonstrated that the B6-38 sequences were inherited with both the X and Y chromosome. B6-38 sequences were genetically mapped as a locus within the PAR using two interspecific backcrosses. The locus defined by B6-38 is designated DXYRp1. Preliminary analyses of recombination between the distal X chromosome gene amelogenin (Amg) and the PAR loci for either TelXY or sex chromosome association (Sxa) suggest that the locus DXYRp1 maps to the distal portion of the PAR.

Sex and the single cell: meiosis in yeast.

Recent studies of Saccharomyces cerevisiae have significantly advanced our understanding of the molecular mechanisms of meiotic chromosome behavior. Structural components of the synaptonemal complex have been identified and studies of mutants defective in synapsis have provided insight into the role of the synaptonemal complex in homolog pairing, genetic recombination, crossover interference, and meiotic chromosome segregation. There is compelling evidence that most or all meiotic recombination events initiate with double-strand breaks. Several intermediates in the double-strand break repair pathway have been characterized and mutants blocked at different steps in the pathway have been identified. With the application of genetic, molecular, cytological, and biochemical methods in a single organism, we can expect an increasingly comprehensive and unified view of the meiotic process.

Genetic determinants of human hypertension.

Hypertension is a common trait of multifactorial determination imparting an increased risk of myocardial infarction, stroke, and end-stage renal disease. The primary determinants of hypertension, as well as the factors which determine specific morbid sequelae, remain unknown in the vast majority of subjects. Knowledge that a large fraction of the interindividual variation in this trait is genetically determined motivates the application of genetic approaches to the identification of these primary determinants. Success in this effort will afford insights into pathophysiology, permit preclinical identification of subjects with specific inherited susceptibility, and provide opportunities to tailor therapy to specific underlying abnormalities. To date, mutations in three genes have been implicated in the pathogenesis of human hypertension: mutations resulting in ectopic expression of aldosterone synthase enzymatic activity cause a mendelian form of hypertension known as glucocorticoid-remediable aldosteronism; mutations in the beta subunit of the amiloride-sensitive epithelial sodium channel cause constitutive activation of this channel and the mendelian form of hypertension known as Liddle syndrome; finally, common variants at the angiotensinogen locus have been implicated in the pathogenesis of essential hypertension in Caucasian subjects, although the nature of the functional variants and their mechanism of action remain uncertain. These early findings demonstrate the feasibility and utility of the application of genetic analysis to dissection of this trait.

Sex proportions of Haemoproteus blood parasites and local mate competition.

Recent genetic evidence suggests that parasitic protozoa often reproduce by "selfing," defined as sexual stages from a single, clonal lineage fertilizing each other. Selfing favors production of an excess of female over male progeny. We tested whether the proportion of male gametocytes of blood parasites of the genus Haemoproteus was affected by variables that could influence the probability of selfing. Proportions of male Haemoproteus gametocytes from 11 passerine host populations were not affected by the age of the parasites' avian hosts, date in season, sex of host, intensity of host's infection, or prevalence of parasites within host populations.