Isolation of Whole Cell Protein Lysates from Mouse Facial Processes and Cultured Palatal Mesenchyme Cells for Phosphoprotein Analysis.
Isolation of Whole Cell Protein Lysates from Mouse Facial Processes and Cultured Palatal Mesenchyme Cells for Phosphoprotein Analysis.
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DOI：
10.3791/63834
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发表时间：
2022-04-01
影响因子：
1.2
通讯作者：
Fantauzzo, Katherine A.
中科院分区：
文献类型：
--
作者：
Rogers, Madison A.;Dennison, Brenna J. C.;Fantauzzo, Katherine A.
哺乳动物颅面的发育是一个复杂的形态学过程，在此过程中，多个细胞群体协调产生额鼻骨架。这些形态学变化是通过不同的信号相互作用启动和维持的，这些信号相互作用通常包括激酶引起的蛋白质磷酸化。在这里，提供了两个例子的生理相关的背景下，在哺乳动物颅面发育过程中的蛋白质磷酸化的研究：小鼠面部的过程中，特别是E11.5上颌突，培养的小鼠胚胎腭间充质细胞来源于E13.5二级腭架。为了克服蛋白质分离过程中去磷酸化的共同障碍，对允许磷蛋白分离的标准实验室方法进行了修改和修改。此外，还提供了在全细胞蛋白裂解物的蛋白质印迹法后正确分析和定量磷蛋白的最佳实践。这些技术，特别是与药理学抑制剂和/或鼠遗传模型相结合，可用于更深入地了解颅面发育过程中各种磷蛋白的动态和作用。
Mammalian craniofacial development is a complex morphological process during which multiple cell populations coordinate to generate the frontonasal skeleton. These morphological changes are initiated and sustained through diverse signaling interactions, which often include protein phosphorylation by kinases. Here, two examples of physiologically-relevant contexts in which to study phosphorylation of proteins during mammalian craniofacial development are provided: mouse facial processes, in particular E11.5 maxillary processes, and cultured mouse embryonic palatal mesenchyme cells derived from E13.5 secondary palatal shelves. To overcome the common barrier of dephosphorylation during protein isolation, adaptations and modifications to standard laboratory methods that allow for isolation of phosphoproteins are discussed. Additionally, best practices are provided for proper analysis and quantification of phosphoproteins following western blotting of whole cell protein lysates. These techniques, particularly in combination with pharmacological inhibitors and/or murine genetic models, can be used to gain greater insight into the dynamics and roles of various phosphoproteins active during craniofacial development.
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影响因子：
2.6
作者：
Ding, H;Wu, XL;Nagy, A
通讯作者：
Nagy, A
影响因子：
3.7
作者：
Fantauzzo KA;Soriano P
通讯作者：
Soriano P
DOI：
10.1073/pnas.79.17.5312
发表时间：
1982-01-01
期刊：
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA-BIOLOGICAL SCIENCES
影响因子：
--
作者：
CHILDS, CB;PROPER, JA;MOSES, HL
通讯作者：
MOSES, HL
影响因子：
4.1
作者：
Bass JJ;Wilkinson DJ;Rankin D;Phillips BE;Szewczyk NJ;Smith K;Atherton PJ
通讯作者：
Atherton PJ
影响因子：
2.7
作者：
Hooper JE;Feng W;Li H;Leach SM;Phang T;Siska C;Jones KL;Spritz RA;Hunter LE;Williams T
通讯作者：
Williams T