Nondisjunctional segregations in Drosophila female meiosis I are preceded by homolog malorientation at metaphase arrest

Abstract

The model of Drosophila female meiosis I was recently revised by the discovery that chromosome congression precedes metaphase I arrest. Use of the prior framework to interpret data from meiotic mutants led to the conclusion that chromosome segregation errors (nondisjunction, NDJ) occurred when nonexchange chromosomes moved out on the spindle in a maloriented configuration and became trapped there at metaphase arrest. The discovery that congression returns nonexchange chromosomes to the metaphase plate invalidates this interpretation and raises the question of what events actually do lead to NDJ. To address this, we have assayed an allelic series of ald (mps1) meiotic mutants that complete congression at wild-type rates, but have widely varying NDJ rates in an otherwise isogenic background, as well as a nod mutant background that primarily undergoes loss of chromosome 4. Using genetic assays to measure NDJ rates, and FISH assays to measure chromosome malorientation rates in metaphase-arrested oocytes, shows that these two rates are highly correlated across ald mutants, suggesting that malorientation during congression commits these chromosomes to eventually nondisjoin. Likewise, the rate of chromosome loss observed in nod is similar to the rate at which these chromosomes fail to associate with the main chromosome mass. Together these results provide a proximal mechanism for how these meiotic mutants cause NDJ and chromosome loss and improve our understanding of how prometaphase chromosome congression relates to anaphase chromosome segregation.

Figure 1

Model for nondisjunctional segregation with proper congression. A heterologous segregation event is drawn here; black arrows indicate chromosome movements. (A) If chromosomes are out on the spindle in a maloriented configuration, and complete congression without correcting the defect, then (B) the metaphase-arrested chromosome mass would contain all the chromosomes, yet will have both homologs pointed toward the same spindle pole. (C) When sister chromatid cohesion along chromosome arms is dissolved at anaphase, the maloriented chromosomes would proceed to opposite poles. If the left pole becomes the pronucleus, the oocyte will be diplo-X nullo-4, while the other pole would produce nullo-X diplo-4.

Figure 2

Immuno-FISH of metaphase arrested oocytes. Oocytes from experimental females from the indicated lines are shown; each was FM7/y w; ald^excision/Df(3R)AN6; pol. For each image, the tubulin spindle (antibody) is shown in gray, DNA (DAPI) in blue, the 4 (FISH probe) in red, and the X (FISH probe) in green. All bars, 4 µm. Small and variable foci of both probes can be found on other chromosomes (Hughes et al. 2009) so the size of the block of probe signal is needed to determine chromosome locations. (A) An ald^excision-14 oocyte showing proper coorientation of both X and 4. FM7 is facing the right pole. (B) An ald^excision-1 oocyte showing proper X coorientation and both 4 chromosomes oriented to the left pole. The left X mass is the distal FM7 signal, with its centromeric spot on top of the combined 4 signal. (C) An ald^excision-4 oocyte showing proper coorientation of both 4 chromosomes with both X chromosomes oriented to the left pole. (D) An ald^excision-23 oocyte showing heterologous segregation, with both 4 chromosomes oriented to the left pole and both X chromosomes oriented to the right. (E) An ald^excision-15 oocyte showing nonheterologous segregation, with all four X and 4 homologs facing the right pole.

Figure 3

Comparison of NDJ and malorientation rates. The rates and 95% confidence intervals in Tables 1 and 2 are plotted for (A) 4 chromosome and (B) X chromosome. The two rates are almost perfectly correlated for both chromosomes (4: r = 0.972, X: r = 0.970, Pearson correlation coefficient) and close to equal (diagonal line).

Figure 4

Chromosome loss in nod mutant oocytes. Experimental virgin females were aged 4 dpe with yeast and no males, and then FISH labeled with probes that predominantly hybridized to the X (green), 4 (red), and 2L–3L (white). As chiasmate chromosomes do not undergo meiotic errors in nod, the two pairs of white 2L–3L foci were used to infer the direction of the meiotic spindles, which are positioned horizontally. Three configurations are shown here, out of 20 possible configurations, not all of which were actually observed (Table S5). All bars, 4 µm. (A) An oocyte from a heterozygous control y w/y w nod^a; pol female, showing biorientation of both X and 4. This oocyte would always produce a euploid oocyte; 200/200 control oocytes scored showed this proper biorientation. (B) An oocyte from a homozygous y w nod^a; pol female, showing the two X homologs correctly bioriented, while the two 4 homologs are together off of the spindle to the right. This X⇔X oocyte would always produce normal-X, nullo-4 progeny. (C) An oocyte from a homozygous y w nod^a; pol female, showing the left pole with no X or 4, and single X and 4 chromosomes associated with the right pole; the other X and 4 homologs were visible elsewhere in the ooplasm (not shown). If the left spindle pole became the egg pronucleus, this Ø⇔X4 oocyte would produce a nullo-X, nullo-4 progeny, while the right pole would result in euploid progeny.