In vitro characterization of the RS motif in N-terminal head domain of goldfish germinal vesicle lamin B3 necessary for phosphorylation of the p34cdc2 target serine by SRPK1.
In vitro characterization of the RS motif in N-terminal head domain of goldfish germinal vesicle lamin B3 necessary for phosphorylation of the p34cdc2 target serine by SRPK1.
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DOI：
10.1016/j.fob.2013.03.003
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发表时间：
2013
期刊：
影响因子：
2.6
通讯作者：
Matsuyama M
中科院分区：
文献类型：
--
作者：
Yamaguchi A;Iwatani M;Ogawa M;Kitano H;Matsuyama M
在低等脊椎动物（鱼和青蛙）的卵母细胞萌发囊泡周围的核膜由核纤层支持，核纤层由核纤层蛋白B3组成，核纤层蛋白B3由一个在鸟类中也发现但在哺乳动物谱系中丢失的基因编码。与核纤层蛋白家族的其他成员一样，金鱼核纤层蛋白B3（gfLB 3）含有两个推定的共有磷酸受体p34 cdc 2位点（Ser-28和Ser-398），用于M期激酶调节核纤层蛋白在中央杆状结构域侧翼的N-和C-末端区域上的聚合。gfLB 3的部分磷酸化发生在未成熟卵母细胞的N-末端头部结构域中的Ser-28上，这表明在鱼类卵母细胞中由一种新的核纤层蛋白激酶引起的核纤层蛋白的连续重排。我们应用表达筛选方法，从金鱼卵巢cDNA文库中分离出磷酸化位点Ser-28特异性单克隆抗体和编码底物肽的载体。因此，SRPK 1被筛选为突出的核纤层蛋白激酶候选者。gfLB 3在N-末端头部内的Ser-28上游具有一段短的RS重复序列（9-SRASTVRSSRRS-20）。这段重复序列在鱼类核纤层蛋白B3中是保守的，但在其他核纤层蛋白中没有发现。体外磷酸化研究和GST下拉试验表明，SRPK 1与连续RS重复序列（9-20）区域具有亲和力，并以抓拉方式将丝氨酸招募到活性位点。这些结果表明SRPK 1可能磷酸化GV-核纤层蛋白B3 N-末端头部的p34 cdc 2位点的RS基序，其具有一般的聚集性质。SRPK 1是从金鱼卵巢中筛选出的一个突出的核纤层蛋白激酶候选者。金鱼核纤层蛋白B3（LB 3）在cdc 2靶位点的上游具有RS重复序列。RS重复序列在鱼类LB 3中是保守的，但在其他核纤层蛋白中没有发现。SRPK 1与RS重复序列亲和结合，并以抓拉方式磷酸化cdc 2位点。
The nuclear envelopes surrounding the oocyte germinal vesicles of lower vertebrates (fish and frog) are supported by the lamina, which consists of the protein lamin B3 encoded by a gene found also in birds but lost in the lineage leading to mammals. Like other members of the lamin family, goldfish lamin B3 (gfLB3) contains two putative consensus phosphoacceptor p34cdc2 sites (Ser-28 and Ser-398) for the M-phase kinase to regulate lamin polymerization on the N- and C-terminal regions flanking a central rod domain. Partial phosphorylation of gfLB3 occurs on Ser-28 in the N-terminal head domain in immature oocytes prior to germinal vesicle breakdown, which suggests continual rearrangement of lamins by a novel lamin kinase in fish oocytes. We applied the expression-screening method to isolate lamin kinases by using phosphorylation site Ser-28-specific monoclonal antibody and a vector encoding substrate peptides from a goldfish ovarian cDNA library. As a result, SRPK1 was screened as a prominent lamin kinase candidate. The gfLB3 has a short stretch of the RS repeats (9-SRASTVRSSRRS-20) upstream of the Ser-28, within the N-terminal head. This stretch of repeats is conserved among fish lamin B3 but is not found in other lamins. In vitro phosphorylation studies and GST-pull down assay revealed that SRPK1 bound to the region of sequential RS repeats (9–20) with affinity and recruited serine into the active site by a grab-and-pull manner. These results indicate SRPK1 may phosphorylate the p34cdc2 site in the N-terminal head of GV-lamin B3 at the RS motifs, which have the general property of aggregation. SRPK1 was screened as a prominent lamin kinase candidate from goldfish ovary. The goldfish lamin B3 (LB3) has RS repeats upstream of the cdc2 target site. The RS repeats are conserved among fish LB3s but are not found in other lamins. SRPK1 binds to the RS repeats with affinity and phosphorylates cdc2 site by a grab-and-pull manner.
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影响因子：
14.9
作者：
Haynes C;Iakoucheva LM
通讯作者：
Iakoucheva LM
影响因子：
4.8
作者：
Lukasiewicz, Randall;Velazquez-Dones, Adolfo;Ghosh, Gourisankar
通讯作者：
Ghosh, Gourisankar
影响因子：
5.6
作者：
Lukasiewicz, Randall;Nolen, Bradley;Ghosh, Gourisankar
通讯作者：
Ghosh, Gourisankar
DOI：
10.1073/pnas.181354098
发表时间：
2001-08-28
影响因子：
11.1
作者：
Lai, MC;Lin, RI;Tarn, WY
通讯作者：
Tarn, WY
影响因子：
3.5
作者：
EGGERT, M;RADOMSKI, N;JOST, E
通讯作者：
JOST, E