Origin of a Giant Sex Chromosome.

Chromosome size and morphology vary within and among species, but little is known about the proximate or ultimate causes of these differences. Cichlid fish species in the tribe Oreochromini share an unusual giant chromosome that is ∼3 times longer than the other chromosomes. This giant chromosome functions as a sex chromosome in some of these species. We test two hypotheses of how this giant sex chromosome may have evolved. The first hypothesis proposes that it evolved by accumulating repetitive elements as recombination was reduced around a dominant sex determination locus, as suggested by canonical models of sex chromosome evolution. An alternative hypothesis is that the giant sex chromosome originated via the fusion of an autosome with a highly repetitive B chromosome, one of which carried a sex determination locus. We test these hypotheses using comparative analysis of chromosome-scale cichlid and teleost genomes. We find that the giant sex chromosome consists of three distinct regions based on patterns of recombination, gene and transposable element content, and synteny to the ancestral autosome. The WZ sex determination locus encompasses the last ∼105 Mb of the 134-Mb giant chromosome. The last 47 Mb of the giant chromosome shares no obvious homology to any ancestral chromosome. Comparisons across 69 teleost genomes reveal that the giant sex chromosome contains unparalleled amounts of endogenous retroviral elements, immunoglobulin genes, and long noncoding RNAs. The results favor the B chromosome fusion hypothesis for the origin of the giant chromosome.

Dynamic Sequence Evolution of a Sex-Associated B Chromosome in Lake Malawi Cichlid Fish.

B chromosomes are extra chromosomes found in many species of plants, animals, and fungi. B chromosomes often manipulate common cellular processes to increase their frequency, sometimes to the detriment of organismal fitness. Here, we characterize B chromosomes in several species of Lake Malawi cichlid fish. Whole genome sequencing of Metriaclima zebra "Boadzulu" individuals revealed blocks of sequence with unusually high sequence coverage, indicative of increased copy number of those sequences. These regions of high sequence coverage were found only in females. SNPs unique to the high copy number sequences permitted the design of specific amplification primers. These primers amplified fragments only in Metriaclima lombardoi individuals that carried a cytologically identified B chromosome (B-carriers), indicating these extra copies are located on the B chromosome. These same primers were used to identify B-carrying individuals in additional species from Lake Malawi. Across 7 species, a total of 43 B-carriers were identified among 323 females. B-carriers were exclusively female; no B chromosomes were observed in the 317 males surveyed from these species. Quantitative analysis of the copy number variation of B-specific sequence blocks suggests that B-carriers possess a single B chromosome, consistent with previous karyotyping of M. lombardoi A single B chromosome in B-carriers is consistent with 2 potential drive mechanisms: one involving nondisjunction and preferential segregation in a mitotic division prior to the germ-line, and the other involving preferential segregation during meiosis I.

An egg-sabotaging mechanism drives non-Mendelian transmission in mice.

Selfish genetic elements drive in meiosis to distort their transmission ratio and increase their representation in gametes, violating Mendel's law of segregation. The two established paradigms for meiotic drive, gamete killing and biased segregation, are fundamentally different. In gamete killing, typically observed with male meiosis, selfish elements sabotage gametes that do not contain them. By contrast, killing is predetermined in female meiosis, and selfish elements bias their segregation to the single surviving gamete (i.e., the egg in animal meiosis). Here, we show that a selfish element on mouse chromosome 2, Responder to drive 2 (R2d2), drives using a hybrid mechanism in female meiosis, incorporating elements of both killing and biased segregation. We propose that if R2d2 is destined for the polar body, it manipulates segregation to sabotage the egg by causing aneuploidy, which is subsequently lethal in the embryo, ensuring that surviving progeny preferentially contain R2d2. In heterozygous females, R2d2 orients randomly on the metaphase spindle but lags during anaphase and preferentially remains in the egg, regardless of its initial orientation. Thus, the egg genotype is either euploid with R2d2 or aneuploid with both homologs of chromosome 2, with only the former generating viable embryos. Consistent with this model, R2d2 heterozygous females produce eggs with increased aneuploidy for chromosome 2, increased embryonic lethality, and increased transmission of R2d2. In contrast to typical gamete killing of sisters produced as daughter cells in a single meiosis, R2d2 prevents production of any viable gametes from meiotic divisions in which it should have been excluded from the egg.

An egg sabotaging mechanism drives non-Mendelian transmission in mice.

During meiosis, homologous chromosomes segregate so that alleles are transmitted equally to haploid gametes, following Mendel's Law of Segregation. However, some selfish genetic elements drive in meiosis to distort the transmission ratio and increase their representation in gametes. The established paradigms for drive are fundamentally different for female vs male meiosis. In male meiosis, selfish elements typically kill gametes that do not contain them. In female meiosis, killing is predetermined, and selfish elements bias their segregation to the single surviving gamete (i.e., the egg in animal meiosis). Here we show that a selfish element on mouse chromosome 2, R2d2, drives using a hybrid mechanism in female meiosis, incorporating elements of both male and female drivers. If R2d2 is destined for the polar body, it manipulates segregation to sabotage the egg by causing aneuploidy that is subsequently lethal in the embryo, so that surviving progeny preferentially contain R2d2. In heterozygous females, R2d2 orients randomly on the metaphase spindle but lags during anaphase and preferentially remains in the egg, regardless of its initial orientation. Thus, the egg genotype is either euploid with R2d2 or aneuploid with both homologs of chromosome 2, with only the former generating viable embryos. Consistent with this model, R2d2 heterozygous females produce eggs with increased aneuploidy for chromosome 2, increased embryonic lethality, and increased transmission of R2d2. In contrast to a male meiotic driver, which kills its sister gametes produced as daughter cells in the same meiosis, R2d2 eliminates "cousins" produced from meioses in which it should have been excluded from the egg.

Unravelling the mystery of female meiotic drive: where we are.

Female meiotic drive is the phenomenon where a selfish genetic element alters chromosome segregation during female meiosis to segregate to the egg and transmit to the next generation more frequently than Mendelian expectation. While several examples of female meiotic drive have been known for many decades, a molecular understanding of the underlying mechanisms has been elusive. Recent advances in this area in several model species prompts a comparative re-examination of these drive systems. In this review, we compare female meiotic drive of several animal and plant species, highlighting pertinent similarities.

Structure and Sequence of the Sex Determining Locus in Two Wild Populations of Nile Tilapia.

In domesticated strains of the Nile tilapia, phenotypic sex has been linked to genetic variants on linkage groups 1, 20 and 23. This diversity of sex-loci might reflect a naturally polymorphic sex determination system in Nile tilapia, or it might be an artefact arising from the process of domestication. Here, we searched for sex-determiners in wild populations from Kpandu, Lake Volta (Ghana-West Africa), and from Lake Koka (Ethiopia-East Africa) that have not been subjected to any genetic manipulation. We analysed lab-reared families using double-digest Restriction Associated DNA sequencing (ddRAD) and analysed wild-caught males and females with pooled whole-genome sequencing (WGS). Strong sex-linked signals were found on LG23 in both populations, and sex-linked signals with LG3 were observed in Kpandu samples. WGS uncovered blocks of high sequence coverage, suggesting the presence of B chromosomes. We confirmed the existence of a tandem amh duplication in LG23 in both populations and determined its breakpoints between the oaz1 and dot1l genes. We found two common deletions of ~5 kb in males and confirmed the presence of both amhY and amh∆Y genes. Males from Lake Koka lack both the previously reported 234 bp deletion and the 5 bp frameshift-insertion that creates a premature stop codon in amh∆Y.

Changing sex for selfish gain: B chromosomes of Lake Malawi cichlid fish.

B chromosomes are extra, non-essential chromosomes present in addition to the normal complement of A chromosomes. Many species of cichlid fish in Lake Malawi carry a haploid, female-restricted B chromosome. Here we show that this B chromosome exhibits drive, with an average transmission rate of 70%. The offspring of B-transmitting females exhibit a strongly female-biased sex ratio. Genotyping of these offspring reveals the B chromosome carries a female sex determiner that is epistatically dominant to an XY system on linkage group 7. We suggest that this sex determiner evolved to enhance the meiotic drive of the B chromosome. This is some of the first evidence that female meiotic drive can lead to the invasion of new sex chromosomes solely to benefit the driver, and not to compensate for skewed sex ratios.

Diurnal variation in opsin expression and common housekeeping genes necessitates comprehensive normalization methods for quantitative real-time PCR analyses.

To determine the visual sensitivities of an organism of interest, quantitative reverse transcription-polymerase chain reaction (qRT-PCR) is often used to quantify expression of the light-sensitive opsins in the retina. While qRT-PCR is an affordable, high-throughput method for measuring expression, it comes with inherent normalization issues that affect the interpretation of results, especially as opsin expression can vary greatly based on developmental stage, light environment or diurnal cycles. We tested for diurnal cycles of opsin expression over a period of 24 hr at 1-hr increments and examined how normalization affects a data set with fluctuating expression levels using qRT-PCR and transcriptome data from the retinae of the cichlid Pelmatolapia mariae. We compared five methods of normalizing opsin expression relative to (a) the average of three stably expressed housekeeping genes (Ube2z, EF1-α and ß-actin), (b) total RNA concentration, (c) GNAT2, (the cone-specific subunit of transducin), (d) total opsin expression and (e) only opsins expressed in the same cone type. Normalizing by proportion of cone type produced the least variation and would be best for removing time-of-day variation. In contrast, normalizing by housekeeping genes produced the highest daily variation in expression and demonstrated that the peak of cone opsin expression was in the late afternoon. A weighted correlation network analysis showed that the expression of different cone opsins follows a very similar daily cycle. With the knowledge of how these normalization methods affect opsin expression data, we make recommendations for designing sampling approaches and quantification methods based upon the scientific question being examined.

Genomic Characterization of a B Chromosome in Lake Malawi Cichlid Fishes.

B chromosomes (Bs) were discovered a century ago, and since then, most studies have focused on describing their distribution and abundance using traditional cytogenetics. Only recently have attempts been made to understand their structure and evolution at the level of DNA sequence. Many questions regarding the origin, structure, function, and evolution of B chromosomes remain unanswered. Here, we identify B chromosome sequences from several species of cichlid fish from Lake Malawi by examining the ratios of DNA sequence coverage in individuals with or without B chromosomes. We examined the efficiency of this method, and compared results using both Illumina and PacBio sequence data. The B chromosome sequences detected in 13 individuals from 7 species were compared to assess the rates of sequence replacement. B-specific sequence common to at least 12 of the 13 datasets were identified as the "Core" B chromosome. The location of B sequence homologs throughout the genome provides further support for theories of B chromosome evolution. Finally, we identified genes and gene fragments located on the B chromosome, some of which may regulate the segregation and maintenance of the B chromosome.