Identification of protein domains required for makorin-2-mediated neurogenesis inhibition in Xenopus embryos.
Makorin-2, consisting of four highly conserved C(3)H zinc fingers, a Cys-His motif and a C(3)HC(4) RING zinc finger domain, is a putative ribonucleoprotein. We have previously reported that Xenopus makorin-2 (mkrn2) is a neurogenesis inhibitor acting upstream of glycogen synthase kinase-3beta (GSK-3beta) in the phosphatidylinositol 3-kinase/Akt pathway. In an effort to identify the functional domains required for its anti-neurogenic activity, we designed and constructed a series of N- and C-terminal truncation mutants of mkrn2. Concurred with the full-length mkrn2, we showed that overexpression of one of the truncation mutants mkrn2(s)-7, which consists of only the third C(3)H zinc finger, Cys-His motif and C(3)HC(4) RING zinc finger, is essential and sufficient to produce the phenotypical dorso-posterior deficiencies and small-head/short-tail phenotype in tadpoles. In animal cap explant assay, we further demonstrated that mkrn2(s)-7 not only inhibits activin and retinoic acid-induced animal cap neuralization and the expression of a pan-neural marker neural cell adhesion molecule, but also induces GSK-3beta expression. These results collectively suggest that the third C(3)H zinc finger, Cys-His motif and C(3)HC(4) RING zinc finger are indispensable for the anti-neurogenic activity of mkrn2.
Cheung
William K C
WK
Department of Chemistry and Open Laboratory of Chemical Biology, The University of Hong Kong, Pokfulam Road, Hong Kong SAR, China.
Yang
Pai-Hao
PH
Huang
Qiu-Hua
QH
Chen
Zhu
Z
Chen
Sai-Juan
SJ
Lin
Marie C M
MC
Kung
Hsiang-Fu
HF
D020816:Amino Acid Motifs / Q000235:genetics; D000818:Animals; D017124:Conserved Sequence; D004625:Embryo, Nonmammalian / Q000378:metabolism; D038362:Glycogen Synthase Kinase 3 / Q000378:metabolism; D055495:Neurogenesis; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism; D010802:Phylogeny; D017434:Protein Structure, Tertiary / Q000235:genetics; D012261:Ribonucleoproteins / Q000145:classification / Q000235:genetics / Q000378:metabolism*; D029867:Xenopus Proteins / Q000145:classification / Q000235:genetics / Q000378:metabolism*; D014982:Xenopus laevis / Q000196:embryology* / Q000235:genetics / Q000378:metabolism; D016335:Zinc Fingers / Q000235:genetics
C528868:Mkrn2 protein, Xenopus; D010750:Phosphoproteins; D012261:Ribonucleoproteins; D029867:Xenopus Proteins; C118206:ZFP36 protein, Xenopus; D038362:Glycogen Synthase Kinase 3