Expression of multiple Src family kinases in sea urchin eggs and their function in Ca2+ release at fertilization.
Expression of multiple Src family kinases in sea urchin eggs and their function in Ca2+ release at fertilization.
复制标题
海胆卵中多种 Src 家族激酶的表达及其在受精时 Ca2 释放中的功能。
DOI：
10.1016/j.ydbio.2008.12.032
复制
发表时间：
2009
影响因子：
2.7
通讯作者：
Foltz,KathyR
中科院分区：
文献类型：
--
作者：
Townley,IanK;Schuyler,Erin;Parker-Gur,Michelle;Foltz,KathyR
后口动物受精时卵子的激活需要细胞内 Ca2+ 的增加，Ca2+ 是从卵子的内质网中释放出来的。在海胆中，Src 家族激酶 (SpSFK1) 对于 PLCγ 介导的信号传导事件是必要的，可启动 Ca2+ 释放（Giusti, A.F.、O'Neill, F.J.、Yamasu, K.、Foltz, K.R. 和 Jaffe, L.A.，2003。海胆卵 Src 家族激酶在受精时启动 Ca2+ 释放的功能。Dev. Biol。 256、367-378。）。对Strongylocentrotus purpuratus基因组序列的注释导致鉴定了额外的预测SFK（Bradham，C.A.，Foltz，D.R.，Beane，W.S.，Amone，M.I.，Rizzo，F.，Coffman，J.A.，Mushegian，A.，Goel，M.，Morales，J.，Geneviere，A.M.，Lapraz， F., Robertson, A.J., Kelkar, H., Loza-Coll, M., Townley, I.K., Raisch, M., Roux, M.M., Lepage, T., Gache, C., McClay, D.R., Manning, G., 2006。海胆激酶组：初看。Dev. 300, 180–193。 M.M.、Townley, I.K.、Raisch, M.、Reade, A.、Bradham, C.、Humphreys, G.、Gunaratne, H.J.、Killian, C.E.、Moy, G.、Su, Y.H.、Ettensohn, C.A.、Wilt, F.、Vacquier, V.D.、Burke, R.D.、Wessel, G. 和 Foltz, K.R.，2006。海胆钙信号传导和卵激活的功能基因组和蛋白质组学视角。300, 416–433。）在这里，我们描述了这 4 个额外 SFK 的克隆和表征，并使用显性干扰显微注射方法结合 Ca2+ 记录测试了它们在受精时初始 Ca2+ 释放过程中的功能。虽然两个新的 SFK（SpFrk 和 SpSFK3）对于 Ca2+ 释放是必需的，但 SpSFK5 对于早期卵到胚胎的转变事件似乎是可有可无的。有趣的是，SpSFK7 可能参与防止 Ca2+ 过早释放。结合研究表明，只有 SpSFK1 能够与 PLCγ 直接相互作用。免疫定位研究表明，一种或多种 SpSFK 和 PLCγ 定位于卵子皮质和精卵相互作用部位。总的来说，这些数据表明不止一种 SFK 参与受精时的 Ca2+ 释放途径。
Egg activation at fertilization in deuterostomes requires a rise in intracellular Ca2+, which is released from the egg's endoplasmic reticulum. In sea urchins, a Src Family Kinase (SpSFK1) is necessary for the PLCγ-mediated signaling event that initiates this Ca2+release (Giusti, A.F., O'Neill, F.J., Yamasu, K., Foltz, K.R. and Jaffe, L.A., 2003. Function of a sea urchin egg Src family kinase in initiating Ca2+ release at fertilization. Dev. Biol. 256, 367–378.). Annotation of the Strongylocentrotus purpuratus genome sequence led to the identification of additional, predicted SFKs (Bradham, C.A., Foltz, D.R., Beane, W.S., Amone, M.I., Rizzo, F., Coffman, J.A., Mushegian, A., Goel, M., Morales, J., Geneviere, A.M., Lapraz, F., Robertson, A.J., Kelkar, H., Loza-Coll, M., Townley, I.K., Raisch, M., Roux, M.M., Lepage, T., Gache, C., McClay, D.R., Manning, G., 2006. The sea urchin kinome: a first look. Dev. Biol. 300, 180–193.; Roux, M.M., Townley, I.K., Raisch, M., Reade, A., Bradham, C., Humphreys, G., Gunaratne, H.J., Killian, C.E., Moy, G., Su, Y.H., Ettensohn, C.A., Wilt, F., Vacquier, V.D., Burke, R.D., Wessel, G. and Foltz, K.R., 2006. A functional genomic and proteomic perspective of sea urchin calcium signaling and egg activation. Dev. Biol. 300, 416–433.). Here, we describe the cloning and characterization of these 4 additional SFKs and test their function during the initial Ca2+release at fertilization using the dominant-interfering microinjection method coupled with Ca2+recording. While two of the new SFKs (SpFrk and SpSFK3) are necessary for Ca2+release, SpSFK5 appears dispensable for early egg to embryo transition events. Interestingly, SpSFK7 may be involved in preventing precocious release of Ca2+. Binding studies indicate that only SpSFK1 is capable of direct interaction with PLCγ. Immunolocalization studies suggest that one or more SpSFK and PLCγ are localized to the egg cortex and at the site of sperm–egg interaction. Collectively, these data indicate that more than one SFK is involved in the Ca2+release pathway at fertilization.