Giroux M J, Hannah L C
Program in Plant Molecular and Cellular Biology, University of Florida, Gainesville 32611.
Mol Gen Genet. 1994 May 25;243(4):400-8. doi: 10.1007/BF00280470.
The Shrunken-2 (Sh2) and Brittle-2 (Bt2) genes of maize encode subunits of the tetrameric maize endosperm ADPglucose pyrophosphorylase. However, in all sh2 and bt2 mutants so far examined, measurable ADPglucose pyrophosphorylase activity remains. We have investigated the origin of the residual activity found in various sh2 and bt2 mutants as well as tissue specific expression and post-translational modification of the Sh2 and Bt2 proteins. Sh2 and Bt2 cDNAs were expressed in Escherichia coli and antibodies were prepared against the resulting proteins SH2 and BT2 specific antibodies were used to demonstrate that SH2 and BT2 are endosperm specific, are altered or missing in various sh2 or bt2 mutants, and have a mol. wt. of 54 and 51 kDa respectively in the wild type. The Sh2 and Bt2 transcripts are also endosperm specific. Ten sh2 and eight bt2 mutants show varying severity of phenotypes expressed at transcript, protein subunit and kernel level. Synthesis of multiple transcripts and proteins commonly occurs as a result of sh2 or bt2 mutation. While all mutants produce detectable enzymic activity, not all produce detectable transcripts and proteins. To examine the origin of the apparent non-SH2/BT2 endosperm enzymic activity, homologs of Sh2 and Bt2, designated Agp1 and Agp2 respectively, were isolated from an embryo cDNA library and found to hybridize to endosperm transcripts distinct from those of Sh2 and Bt2. Thus Agp1 and Agp2 or closely related genes may be responsible for the residual activity in some sh2 and bt2 mutants. Surprisingly, no evidence of post-translational modification of the SH2 and BT2 protein subunits was detected.
玉米的皱缩-2（Sh2）基因和脆-2（Bt2）基因编码四聚体玉米胚乳ADP葡萄糖焦磷酸化酶的亚基。然而，在目前检测的所有sh2和bt2突变体中，仍存在可测量的ADP葡萄糖焦磷酸化酶活性。我们研究了各种sh2和bt2突变体中残余活性的来源，以及Sh2和Bt2蛋白的组织特异性表达和翻译后修饰。Sh2和Bt2 cDNA在大肠杆菌中表达，并制备了针对所得蛋白的抗体。使用SH2和BT2特异性抗体证明，SH2和BT2是胚乳特异性的，在各种sh2或bt2突变体中发生改变或缺失，并且在野生型中分子量分别为54 kDa和51 kDa。Sh2和Bt2转录本也是胚乳特异性的。十个sh2突变体和八个bt2突变体在转录本、蛋白质亚基和籽粒水平上表现出不同严重程度的表型。sh2或bt2突变通常会导致多种转录本和蛋白质的合成。虽然所有突变体都产生可检测的酶活性，但并非所有突变体都产生可检测的转录本和蛋白质。为了研究明显的非SH2/BT2胚乳酶活性的来源，分别从胚胎cDNA文库中分离出Sh2和Bt2的同源物，命名为Agp1和Agp2，发现它们与不同于Sh2和Bt2的胚乳转录本杂交。因此，Agp1和Agp2或密切相关的基因可能是一些sh2和bt2突变体中残余活性的原因。令人惊讶的是，未检测到SH2和BT2蛋白亚基翻译后修饰的证据。
