Kofron M et al. (1997), The roles of maternal alpha-catenin and plakogl...

XB-ART-16698

Development 1997 Apr 01;1248:1553-60. doi: 10.1242/dev.124.8.1553.

Show Gene links Show Anatomy links

The roles of maternal alpha-catenin and plakoglobin in the early Xenopus embryo.

Kofron M , Spagnuolo A , Klymkowsky M , Wylie C , Heasman J .

???displayArticle.abstract???
Catenins (alpha-, beta- and gamma- or plakoglobin) are cytoplasmic cadherin-associated proteins. Studies on cultured cells have suggested that both alpha-catenin and plakoglobin are important for the adhesive function of cadherins. alpha-catenin binds to both beta-catenin and plakoglobin and may link the cadherin/catenin complex to actin filaments. Separate domains of plakoglobin bind to cadherin and alpha-catenin, suggesting it may act as a bridge between these molecules. However, plakoglobin may have other activities: it is expressed in both desmosomal junctions in association with desmogleins and the cytoplasm in conjunction with APC, and previous work suggests it may act in a dorsal signalling pathway when overexpressed in Xenopus embryos. Here, we have studied the roles of alpha-catenin and plakoglobin directly, by depleting the maternal mRNAs coding for each of them in developing Xenopus embryos. We find that depletion of maternal alpha-catenin causes the loss of intercellular adhesion at the blastula stage, similar to that reported previously for EP cadherin. Depletion of plakoglobin results in a partial loss of adhesion, and a loss of embryonic shape, but does not affect dorsal signalling.

???displayArticle.pubmedLink??? 9108371
???displayArticle.link??? Development
???displayArticle.grants??? [+]

Species referenced: Xenopus laevis
Genes referenced: actl6a ctnnb1 jup myod1 tbx2

???attribute.lit??? ???displayArticles.show???

	Fig. 1. Injection of antisense oligos complementary to a-catenin (A,C,E) or plakoglobin (B,D,E) mRNA depletes the endogenous maternal mRNAs and proteins. (A) Northern blot, probed for a-catenin, of RNA (2 oocytes or embryos equivalent per lane) from cleavage stage (stage 4), mid-blastula stage (stage 8), gastrula stage (stage 10.5) and late neurula stage (stage 18), either uninjected (U) or injected as oocytes with 4 ng oligo A3 (A), and with 4 ng of oligo A3 plus 1.2 ng of mouse a-catenin mRNA (A+M). The exogenous mRNA is smaller because it contains only the mouse a-catenin coding sequence. (B) Northern blot, probed for plakoglobin, of RNA from oocytes, midblastula stage (stage 8), gastrula stage (stage 10.5), late neurula stage (stage 18), either uninjected (U) or injected as oocytes with 2 ng of oligo M7 (A), and with 2 ng of oligo M7 plus 2 ng of plakoglobin mRNA. Both blots were stripped and reprobed for EF1aRNA, to indicate RNA loading. (C,D) Western blots of proteins from stage 6 oocytes (C) and blastula-stage embryos (D), either uninjected or injected as oocytes with 4 ng of oligo A3 (C) or with 2.5 and 3 ng of oligo M7 (D). The blots were probed with a-catenin (C) and plakoglobin (D) antibodies (see Materials and Methods). (E,F) Western blots of supernatant (S) or insoluble (P) fractions of proteins from blastula (stage 8) embryos (5 embryos per fractionation). (E) Lefthand four lanes, embryos injected with 2.5 ng of oligo M7 and probed with plakoglobin antibody, compared to uninjected controls; right-hand four lanes uninjected embryos compared to embryos injected with 5 ng of oligo A3 and probed with a-catenin antibody. (F) Left-hand six lanes, control embryos compared to those injected with 1 or 1.2 ng of oligo 303 (Heasman et al., 1994b) and probed with b-catenin antibody; righthand two lanes, control embryos probed for b-1 integrin.
	Fig. 2. Depletion of maternal a-catenin mRNA and protein causes a disaggregated phenotype, which can be rescued using mouse a-catenin mRNA. (A) A control embryo (left) together with an a-catenin-depleted sibling embryo (right) dissected at the blastula stage to show the loss of interblastomere adhesion. (B) Midblastulae disaggregated and allowed to aggregate for 45 minutes. Blastomeres from a-catenin-deficient embryos (right) failed to aggregate in comparison to wild-type blastomeres (left). (C) Blastulae derived from uninjected (middle) or injected as oocytes with 4 ng oligo A3 (right), and 4 ng oligo A3 plus 1 ng mouse a-catenin mRNA (left) to show that mouse a-catenin mRNA was able to restore a normal blastula morphology into a-catenin-depleted embryos.
	Fig. 3. Depletion of maternal plakoglobin mRNA and protein causes a phenotype marked by a specific affect on cytoarchitecture, a mild adhesion defect most evident in the vegetal blastomeres and a delay of gastrulation. All aspects of this phenotype are at least partially rescued with Xenopus plakoglobin mRNA. (A) A control embryo (top left, brown), a plakoglobin mRNA-treated embryo (bottom left, red) and two oligo doses of plakoglobin-depleted embryos, 2.0 ng (top right, blue) and 2.5 ng (bottom right, mauve). (B) Midblastulae disaggregated and allowed to aggregate for 45 minutes. Blastomeres from plakoglobin-depleted embryos (right) aggregate less well than control blastomeres (left). (C) Comparison of devitellined plakoglobin-depleted (left, mauve) embryos to a-catenin-depleted embryos (right, red). Plakoglobin-depleted embryos show a flattened morphology that is not evident in the a- catenin-depleted embryos. (D) Plakoglobindepleted embryos are delayed in gastrulation. A control embryo (top left, brown), a plakoglobin mRNA-treated embryo (top right, red), and three doses of plakoglobin oligo M7 (bottom left to right) 2.5 ng (mauve), 2.0 ng (blue), 1.75 ng (green). (E) Control and plakoglobin-depleted embryos form normal neural structures. Starting from left to right, plakoglobin-depleted embryos 2.5 ng (mauve), 2.0 ng (blue), 1.75 ng (green), control embryo (brown) and plakoglobin mRNA-treated embryo (red). (F) Partial rescue of oligo-mediated adhesion and cytoarchitectural defects at blastula stage with plakoglobin mRNA. Control embryos (top), plakoglobin-depleted embryos (middle) and plakoglobin-depleted embryos injected with Xenopus plakoglobin mRNA (bottom). (G) Plakoglobin mRNA can rescue the gastrulation delay of plakoglobin-depleted embryos. Plakoglobin-depleted embryos (left),depleted embryos injected with Xenopus plakoglobin mRNA (middle) and control embryos (right).
	Fig. 4. Northern blot probed for MyoD, a marker of dorsal mesoderm. One embryo equivalent was loaded per lane at the gastrula or neurula stage. Experimental embryos were derived from oocytes injected with 1.75 ng, 2.0 ng or 2.5 ng of plakoglobin oligo M7. The blot was stripped and reprobed with Ef1-ato control for loading.