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. Author manuscript; available in PMC: 2009 Mar 2.
Published in final edited form as: Dev Cell. 2005 Sep;9(3):339–349. doi: 10.1016/j.devcel.2005.06.009

Figure 5.

Figure 5

sea-1 Activates xol-1 Transcription

(A-C) Confocal images of wild-type, sea-1 mutant, and sea-1(+) overexpressing embryos at approximately the 20- to 30-cell stage, costained with antibodies against SEA-1 (green) and SDC-3 (red) and with DAPI (blue), a DNA-intercalating dye.

(A) SEA-1 accumulates in nuclei of 20- to 100-cell wild-type embryos, the correct time to regulate xol-1. At the 30-cell stage, the dosage compensation protein SDC-3 is present in nuclei but has not yet localized to the X chromosomes.

(B) SEA-1 protein is not detectable in sea-1(y356) mutant embryos; SDC-3 is unaffected.

(C) sea-1 is overexpressed in yIs61[sea-1(+)] embryos.

(D) sea-1 acts upstream of xol-1. Multiple copies of sea-1 on the integrated array yIs61 enhanced the weak XX-specific lethality caused by sdc-2(RNAi). The enhancement of lethality was suppressed by a xol-1 mutation. n is the total number of embryos counted. Percent viability is the number of adults relative to the number of embryos laid.

(E) Model for sea-1 action. Our cumulative data establish sea-1 to be an ASE that acts upstream in the sex determination hierarchy to activate xol-1 transcription.

(F) DIC images of gravid wild-type XX hermaphrodites, him-8 XX hermaphrodites, and SEA-1-overexpressing XX hermaphrodites carrying yIs33[Pxol-l::lacZ], a transcriptional fusion between the xol-1 promoter and lacZ coding sequence, stained for β-galactosidase activity (Nicoll et al., 1997). Expression of this reporter mimics endogenous xol-1 expression: β-galactosidase is produced in XO embryos but not XX embryos. The him-8 mutation elevates X chromosome nondisjunction, resulting in ∼37% XO progeny compared to 0.2% XO progeny from wild-type XX hermaphrodites. In 58% of yIs33[Pxol-l::lacZ]; him-8(-) hermaphrodites examined, at least one embryo showed dark staining, indicating expression of Pxol-1::lacZ at high level (n = 53 adults containing 590 embryos). These darkly staining embryos are XO. The vast majority of all other embryos (XX) showed no β-galactosidase staining. By contrast, in 87% of gravid yIs33[Pxol-l::lacZ]; him-8(+) XX hermaphrodites examined, more than 75% of the embryos showed either light or no β-galactosidase staining (n = 69 adults containing 1260 embryos). None of the yIs33[Pxol-l::lacZ] hermaphrodites had any darkly staining embryos. Overexpression of sea-1 by the integrated yIs61[sea-1(+)] array was sufficient to partially activate Pxol-1::lacZ expression in XX embryos. About 58% of yIs61[sea-1(+)]; yIs33[Pxol-l::lacZ] gravid hermaphrodites contained at least one darkly staining XX embryo, and 25% of the other XX embryos showed medium to dark staining (n = 62 adults containing 917 embryos). Scale bars equal 5 μm.