Tessmar K et al. (2002), A screen for co-factors of Six3.

XB-ART-6624

Mech Dev 2002 Sep 01;1171-2:103-13. doi: 10.1016/s0925-4773(02)00185-5.

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A screen for co-factors of Six3.

Tessmar K , Loosli F , Wittbrodt J .

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The vertebrate Six3 gene a homeobox gene of the Six-family, plays a crucial role in early eye and forebrain development. Here we report the isolation of candidate factors that interact with Six3 in a yeast two-hybrid screen. Among these are two basic helix loop helix (bHLH) domain containing proteins. Biochemical analysis reveals that the bHLH proteins ATH5, ATH3, NEUROD as well as ASH1 interact specifically with XSix3. By defining the interacting domains we show that the bHLH domain of NEUROD interacts with the SIX domain of XSix3. The co-expression of the interacting molecules during late retina determination/differentiation suggests a new role for Six3 and the respective interaction partner also in these late steps of eye development.

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Species referenced: Xenopus laevis
Genes referenced: actl6a arnt ascl1 atoh7 atp5if1 bmal2 chmp1a cldn6.2 neurod1 neurod4 scoc six3 six6 smarcc1 strn tacc3 tle2 znf326

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	Fig. 1. Comparison of the expression domains of XSix3 and its potential interacting partners. Whole-mount in situ analysis at different developmental stages, lateral views; anterior is to the left. Clone names are indicated in the lower left corner of each picture, and stages in the lower right corner. (A) XSix3 expression at stage 17 (inset in (A)), 21 (A) 24 (B) and 37. (D) The first two columns show candidate interactors expressed at early and later stages; staining was performed at stages 23/24 and at 17 (small insets). Note that all these genes, except for XclaudinA (I), are expressed in the prospective eye and forebrain region from stage 17 onwards. (K) The third column depicts the expression of genes that are co-expressed with XSix3 only later in development (stage 24 or later). Expression domains are shown at stages 37 and 24 or 28 (small insets).
	Fig. 5. Co-expression of XSix3 and XneuroD. Double whole-mount in situ hybridisation detecting XSix3 (blue) and Xath5 (red, A) and XNeuroD (red, D). Anterior to the left, dorsal (A, C, D, G, J, M), lateral (B, E, H, K, N) and frontal views (F, I). Stages as indicated. (A) Full overlap of expression Xath5 and XSix3 detectable only in embryos weakly stained for XSix3. Compare A and J for XSix3. (D) No overlap of expression detected at stage 21. (G) Coexpression first detectable at stage 24. Note that at this stage only scattered individual cells express XNeuroD in a broad XSix3 domain (arrow in inset in H). In addition, the presumptive pineal organ expresses both genes (arrowhead in H and K, compare this to Fig. 1B). (J) At stage 28, co-expression broadens, comprising almost the entire retina (arrowhead in L). (M) At stage 37, co-expression becomes restricted to marginal regions (arrowheads in O). (L, O) Transversal sections at levels indicated in J and M, dorsal to the left, inset shows NeuroD single staining.
	bmal2 (basic helix-loop-helix ARNT like 2) gene expression in Xenopus laevis embryos, NF stage 23 (stage 17 inset), as assayed by in situ hybridization, Lateral view, anterior left, dorsal up.
	chmp1a (charged multivesicular body protein 1A) gene expression in Xenopus laevis embryos, NF stage 37 (stage 24 inset), as assayed by in situ hybridization. Lateral view: anterior left, dorsal up.
	cldn6.2 (claudin 6, gene 2) gene expression in Xenopus laevis embryos, NF stage 24 (stage 17 inset), as assayed by in situ hybridization. Lateral view: anterior left, dorsal up.
	scoc (short coiled-coil protein) gene expression in Xenopus laevis embryos, NF stage 37 (stage 24 inset), as assayed by in situ hybridization, lateral view, anterior left, dorsal up.
	smarcc1 (SWI/SNF related, matrix associated, actin dependent regulator of chromatin, subfamily c, member 1) gene expression in Xenopus laevis embryos, NF stage 23 (stage 17 inset), as assayed by in situ hybridization. Lateral view: anterior left, dorsal up.
	strn (striatin, calmodulin binding protein) gene expression in Xenopus laevis embryos, NF stage 37 (stage 24 inset), as assayed by in situ hybridization. Lateral view: anterior left, dorsal up.
	tacc3 (transforming, acidic coiled-coil containing protein 3 ) gene expression in Xenopus laevis embryos, NF stage 24 (stage 17 inset), as assayed by in situ hybridization. Lateral view: anterior left, dorsal up.