CHARACTERIZATION OF QUAIL PAX-6 (PAX-QNR) PROTEINS EXPRESSED IN THE NEURORETINA
CHARACTERIZATION OF QUAIL PAX-6 (PAX-QNR) PROTEINS EXPRESSED IN THE NEURORETINA
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DOI：
10.1128/mcb.13.12.7257
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发表时间：
1993-12-01
影响因子：
5.3
通讯作者：
SAULE, S
中科院分区：
文献类型：
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作者：
CARRIERE, C;PLAZA, S;SAULE, S
从感染v-myc的禽类逆转录病毒MC29感染的鹌鹑视网膜神经细胞(QNR)中筛选出一个与小鼠常染色体显性突变小眼和人类无虹膜疾病中突变的小鼠Pax-6同源的cDNA克隆Pax-QNR。在这里，我们报告了在鸟类神经视网膜中表达的Pax-QNR蛋白的特征。从细菌表达的Pax-QNR多肽中，我们获得了针对该蛋白不同结构域的兔抗血清：配对结构域(血清11)、配对结构域与同源结构域之间的结构域(血清12)、同源结构域(血清13)和羧基末端部分(血清14)。血清12、13和14能够特异性识别神经视网膜中的5种蛋白质(48、46、43、33和32 kDa)。与48 kDa、46 kDa和43 kDa的蛋白质相比，33 kDa和32 kDa的蛋白质不能被配对的抗血清(血清11)识别。无配对和含配对的蛋白质具有相同的半衰期(6h)，并且主要在丝氨酸残基上被磷酸化。免疫沉淀法显示，成对的蛋白位于胞核，而33和32 kDa的蛋白主要位于细胞质中。然而，瞬时转染后进行的免疫荧光实验表明，p46和p33/32也在体内定位于细胞核内。因此，Pax-QNR/Pax-6基因可以产生具有两个DNA结合域的蛋白质以及只含有DNA结合同源结构域的蛋白质。
After differential screening of a cDNA library constructed from quail neuroretina cells (QNR) infected with the v-myc-containing avian retrovirus MC29, we have isolated a cDNA clone, Pax-QNR, homologous to the murine Pax-6, which is mutated in the autosomal dominant mutation small eye of mice and in the disorder aniridia in humans. Here we report the characterization of the Pax-QNR proteins expressed in the avian neuroretina. From bacterially expressed Pax-QNR peptides, we obtained rabbit antisera directed against different domains of the protein: paired domain (serum 11), domain between the paired domain and homeodomain (serum 12), homeodomain (serum 13), and carboxyl-terminal part (serum 14). Sera 12, 13, and 14 were able to specifically recognize five proteins (48, 46, 43, 33, and 32 kDa) in the neuroretina. In contrast to proteins of 48, 46, and 43 kDa, proteins of 33 and 32 kDa were not recognized by the paired antiserum (serum 11). Paired-less and paired-containing proteins exhibited the same half-life (6 h) and were phosphorylated mostly on serine residues. Immunoprecipitations performed with subcellular fractions of neuroretinas showed that the paired-containing proteins were located in the nucleus, whereas the 33- and 32-kDa proteins were found essentially in the cytoplasmic compartment. However, immunofluorescence experiments performed after transient transfections showed that p46 and p33/32 were also located in vivo into the nucleus. Thus, the Pax-QNR/Pax-6 gene can produce proteins with two DNA-binding domains as well as proteins containing only the DNA-binding homeodomain.