[Effects of site-directed mutagenesis at amino acid residues of GATA-1b different from that of GATA-1a in Xenopus].
The GATA-1 of Xenopus (xGATA-1), which has two subtypes xGATA-1a and xGATA-1b, is a necessary factor for erythroid differentiation and maturation as similar as that of other GATA-1s. Although both xGATa-1a and xGATA-1b are able to stimulate erythropoiesis, only xGATA-1b is capable of inhibiting neurogenesis in Xenopus embryos. Compared between their structures, xGATA-1a and xGATA-1b are very similar in nucleotide and amino acids composition, but not identical. Therefore, it is responsible for studying the role of the diverse codons between the two genes, so the desired mutations: S(168), H(169) double deletion and point mutation of T(304)-->A, T(359)-->A, were introduced into xGATA-1b gene through site-directed mutagenesis. Then, mRNA from each mutant as well as wtxGATA-1b was co-injected with DN-BR mRNA or separately injected into Xenopus stage 2 embryos, and the role of mutants in erythropoiesis and neurogenesis was analyzed by using animal cap culture system. The results showed that the neural-inhibiting activity of xGATA-1b, but not hematopoiesis-inducing activity, was aborted because of deletion of Ser(168) and His(169) or point mutation of T(359)-->A. So it is demonstrated for the first time that Ser(168) and His(169) or Thr(359)in xGATA-1b may be one of the structural basis for explanting the different function between xGATA-1b and xGATA-1a.
Feng
Xiang-Ling
XL
Cancer Research Institute, Xiangya Medical School of Central South University, Changsha 410078, China.
Liu
Wei-Dong
WD
Li
Hong
H
Wang
Lei
L
Yang
Xu-Yu
XY
Han
Wei-Nong
WN
Zhou
Wen
W
Yao
Kai-Tai
KT
D000596:Amino Acids / Q000235:genetics*; D000818:Animals; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000502:physiology; D004625:Embryo, Nonmammalian / Q000187:drug effects / Q000378:metabolism; D047108:Embryonic Development; D050981:Erythroid-Specific DNA-Binding Factors; D004920:Erythropoiesis / Q000502:physiology; D006639:Histidine / Q000235:genetics; D007267:Injections; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D009420:Nervous System / Q000196:embryology; D020033:Protein Isoforms / Q000235:genetics / Q000502:physiology; D012333:RNA, Messenger / Q000008:administration & dosage / Q000235:genetics / Q000378:metabolism; D012694:Serine / Q000235:genetics; D013912:Threonine / Q000235:genetics; D014157:Transcription Factors / Q000235:genetics* / Q000502:physiology; D014982:Xenopus laevis
D000596:Amino Acids; D004268:DNA-Binding Proteins; D050981:Erythroid-Specific DNA-Binding Factors; D020033:Protein Isoforms; D012333:RNA, Messenger; D014157:Transcription Factors; D013912:Threonine; D012694:Serine; D006639:Histidine