Identification of ORD, a Drosophila protein essential for sister chromatid cohesion.

Attachment between the sister chromatids is required for proper chromosome segregation in meiosis and mitosis, but its molecular basis is not understood. Mutations in the Drosophila ord gene result in premature sister chromatid separation in meiosis, indicating that the product of this gene is necessary for sister chromatid cohesion. We isolated the ord gene and found that it encodes a novel 55 kDa protein. Some of the ord mutations exhibit unusual complementation properties, termed negative complementation, in which particular alleles poison the activity of another allele. Negative complementation predicts that protein-protein interactions are critical for ORD function. The position and nature of these unusual ord mutations demonstrate that the C-terminal half of ORD is essential for sister chromatid cohesion and suggest that it mediates protein binding.

Double or nothing: a Drosophila mutation affecting meiotic chromosome segregation in both females and males.

We describe a Drosophila mutation, Double or nothing (Dub), that causes meiotic nondisjunction in a conditional, dominant manner. Previously isolated mutations in Drosophila specifically affect meiosis either in females or males, with the exception of the mei-S332 and ord genes which are required for proper sister-chromatid cohesion. Dub is unusual in that it causes aberrant chromosome segregation almost exclusively in meiosis I in both sexes. In Dub mutant females both nonexchange and exchange chromosomes undergo nondisjunction, but the effect of Dub on nonexchange chromosomes is more pronounced. Dub reduces recombination levels slightly. Multiple nondisjoined chromosomes frequently cosegregate to the same pole. Dub results in nondisjunction of all chromosomes in meiosis I of males, although the levels are lower than in females. When homozygous, Dub is a conditional lethal allele and exhibits phenotypes consistent with cell death.

Sister-chromatid misbehavior in Drosophila ord mutants.

In Drosophila males and females mutant for the ord gene, sister chromatids prematurely disjoin in meiosis. We have isolated five new alleles of ord and analyzed them both as homozygotes and in trans to deficiencies for the locus, and we show that ord function is necessary early in meiosis of both sexes. Strong ord alleles result in chromosome nondisjunction in meiosis I that appears to be the consequence of precocious separation of the sister chromatids followed by their random segregation. Cytological analysis in males confirmed that precocious disjunction of the sister chromatids occurs in prometaphase I. This is in contrast to Drosophila mei-S332 mutants, in which precocious sister-chromatid separation also occurs, but not until late in anaphase I. All three of the new female fertile ord alleles reduce recombination, suggesting they affect homolog association as well as sister-chromatid cohesion. In addition to the effect of ord mutations on meiosis, we find that in ord2 mutants chromosome segregation is aberrant in the mitotic divisions that produce the spermatocytes. The strongest ord alleles, ord2 and ord5, appear to cause defects in germline divisions in the female. These alleles are female sterile and produce egg chambers with altered nurse cell number, size, and nuclear morphology. In contrast to the effects of ord mutations on germline mitosis, all of the alleles are fully viable even when in trans to a deficiency, and thus exhibit no essential role in somatic mitosis. The ord gene product may prevent premature sister-chromatid separation by promoting cohesion of the sister chromatids in a structural or regulatory manner.

The Drosophila mei-S332 gene promotes sister-chromatid cohesion in meiosis following kinetochore differentiation.

The Drosophila mei-S332 gene acts to maintain sister-chromatid cohesion before anaphase II of meiosis in both males and females. By isolating and analyzing seven new alleles and a deficiency uncovering the mei-S332 gene we have demonstrated that the onset of the requirement for mei-S332 is not until late anaphase I. All of our alleles result primarily in equational (meiosis II) nondisjunction with low amounts of reductional (meiosis I) nondisjunction. Cytological analysis revealed that sister chromatids frequently separate in late anaphase I in these mutants. Since the sister chromatids remain associated until late in the first division, chromosomes segregate normally during meiosis I, and the genetic consequences of premature sister-chromatid dissociation are seen as nondisjunction in meiosis II. The late onset of mei-S332 action demonstrated by the mutations was not a consequence of residual gene function because two strong, and possibly null, alleles give predominantly equational nondisjunction both as homozygotes and in trans to a deficiency. mei-S332 is not required until after metaphase I, when the kinetochore differentiates from a single hemispherical kinetochore jointly organized by the sister chromatids into two distinct sister kinetochores. Therefore, we propose that the mei-S322 product acts to hold the doubled kinetochore together until anaphase II. All of the alleles are fully viable when in trans to a deficiency, thus mei-S332 is not essential for mitosis. Four of the alleles show an unexpected sex specificity.