The dice of fate: the csd gene and how its allelic composition regulates sexual development in the honey bee, Apis mellifera

Introduction

A diagram depicting how haplodiploid sex determination occurs in honey bees. (3)

The mechanisms that underlie sex determination appear to share some general features in all species. There is a cascade of genes required to produce male and female phenotypes, however, the initial signals of sex determination differ among species. In Apis mellifera, sex is determined by the fertilization or non-fertilization of the egg rather than the presence or absence of sex chromosomes. The term haplodiploidy refers to a case in which unfertilized haploid eggs develop into males and fertilized diploid eggs develop into females. However, researchers have found that this haplodiploid sex determination may not be the only mechanism at work when determining sex in honey bees.

Complementary sex determination: a history of 150 years of research

Genotypes and sexual fate under the system of complementary sex determination found in many hymenopteran species. Males derive from unfertilized eggs and have only one sex determination allele (marked by different colored bars). Fertilized eggs with two different sex-determining alleles (heterozygous) develop into females. Diploid males arise from fertilized eggs that are homozygous for the same sex-determining allele. These diploid males arise most commonly under inbreeding conditions in which the father has an allele in common with the mother (2).

In 1845, Johann Dzierzon described how male honey bees develop from unfertilized eggs, based on a report that a non mated queen honey bee can only produce males. Cytological studies showed that designated male eggs are not fertilized and are genetically haploid, while eggs that develop into females are diploid. This mode of sex determination occurs in about 20% of all known animal species (4). In the years following Dzierzon‘s discovery, investigators performed inbreeding studies, and were surprised to find that diploid males appeared (2). Since the appearance of diploid males was associated with inbreeding, investigators proposed a hypothesis of complementary sex determination, in which a single sex determination locus (SDL) determines the sexual fate of individuals (2). According to this hypothesis, fertilized eggs that are homozygous at the SDL develop into diploid males, while fertilized eggs that are heterozygous at SDL develop into females. Homozygosity at the SDL is lethal to males. The eggs that differentiate into diploid males are sterile and are eaten by worker bees shortly after the larvae hatch from the egg (4).

Positional cloning identifies the complementary sex determiner

Beye and colleagues used a positional cloning and fine-scale mapping approach to isolate and identify the genomic region of the complimentary sex determiner (csd). The natural occurring trait of females and diploid males in an inbred cross were used to establish markers that were co-segregating with diploid male and female development (1). Two sex-linked markers (Q and Z) were identified from two different labs that joined forces and found that the two markers were flanking the sex-determining region at a distance of less than 360 kb (1). There was a 12 kb region between two genetic markers identified as always heterozygous in females. Exon prediction algorithms and subsequent analysis of transcripts identified a single gene in the sequence of the designated region, which was named the complimentary sex determiner (csd) (1). Studies on developing eggs showed that in both males and females, the csd gene becomes active about 12 hours after eggs are laid and remains active throughout development (4).

Schematic presentation of the genomic organization and proposed protein structure of csd. A: Genomicregionencompassing csd and the two closely linked genetic markers (marker 1 and 2). The genetic markers 1 and 2 are always heterozygous in females; this identifies the sex-determining locus. Exons are indicated by boxes and the deduced ORF is marked in yellow. The estimate of the transcript is indicated in nucleotides (nt), which represents just one allelic variant. No sex-specific differences were found among the transcripts. Predicted translational start and stop sites are indicated. B: The schematic presentation of the predicted domain structure of csd. The region rich in arginine/serine (RS domain) is marked in red, the hypervariable region that has different number of repeats in various alleles is shown in grey (HV) and the proline-rich region is marked in blue. The number of amino acids of the CSD protein varies between alleles as indicated (1).

Csd consists of nine exons and spans a genomic region of about 9 kb. Transcripts are the same in males and females. Repression of csd by RNAi in females that were heterozygous for csd resulted in a full developmental switch into male gonad development (2). Repression of csd in males had no phenotypic effect. When derived from different alleles, csd is functionally active and initiates female development. When derived from one allele, csd is not functionally active and the default male development ensues (2).

Sex-determining alleles and proposed molecular function

Models for protein association of two csd allelic polypeptides and their functional consequences (1).

The single amino acid differences found among the sequences are a potential source of allelic specificity. There are three models to explore how the different specificities can combine and form an active or a non-active molecule. These are heterodimer, homodimer, complementation (1). In heterodimer and homodimer models, binding differences exist depending on whether polypeptides combine from the same or different alleles. The third model, complementation, shows occurrences in activity differences without distinction in binding (1).

Results

In the honey bee, the initial signal depends on whether the allelic composition is identical or different. All alleles have the same potential to initiate male and female development (1). Csd thus defines a class of initiating primary signals in which the genetic composition of the gamete has no predictive value for sex tendency prior to fertilization. However, there is no evidence so far for a mechanism that maintains the sexual fate throughout development (1). One hypothesis is that the permanent transcription of csd, which starts after cell formation, is a constant source of sexual identity (4).

Future

Scientists are starting to understand how the decision for sex determination is made at the level of polypeptides. Comparing the structural and possible functional similarity of the initial signal csd in Apis mellifera and sex determining genes of other insects, offers the opportunity to elucidate how different sex-determining systems have evolved. Further studies will show whether csd is involved in alternative splicing and whether csd directly targets primary transcripts (1).

Strengths and Weaknesses

There was a lot of information on the study that Martin Beye and his colleagues performed. The figures were easy to understand, and assisted with the understanding of the experiment and findings. He wrote briefly about comparing the csd gene with the sex determining genes of Drosophila melanogaster, however, I feel that a more in depth comparison of the two species would have been helpful in understanding the significance of the discovery of the csd gene.

References

1. Beye, Martin. “The Dice of Fate: The Csd Gene and How Its Allelic Composition Regulates Sexual Development in the Honey Bee, Apis Mellifera.” BioEssays26.10 (2004): 1131-139. Print.

2. Gempe, T. & Beye, M. (2009) Sex determination in honeybees. Nature Education 2(2)

3. Haplodiploidy in bees. Art. Encyclopædia Britannica Online.

4. Page, Robert, Martin Beye, Andy Fell, and Pat Bailey. UC Davis News & Information :: Honeybee Gene Find Ends 150-year SearchHoneybee Gene Find Ends 150-year Search. UC Davis News & Information, 21 Aug. 2003.

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