Protein kinases in growth and differentiation of intestinal epithelium

Date of Award




Degree Name

Doctor of Philosophy (Ph.D.)

First Committee Member

David R. Burgess - Committee Chair


Undifferentiated, highly mitotic cells of the crypt small intestine display 15-fold higher cytoskeletal-associated tyrosine kinase activity than cytoskeletons of differentiated villus tip cells. Two of the most abundant tyrosyl phosphorylated proteins associated with the crypt cytoskeletons have M$\sb{\rm r}$ of 36 and 17 kD.One of the cytoskeletal-associated protein tyrosine kinases identified in both crypt and villus tip cells is the protein product of the protooncogene pp60$\sp{c-src}$. Immune complex assays of pp60$\sp{c-src}$ isolated from crypt and villus tip cell cytoskeletons show that the protein tyrosine kinase activity of pp60$\sp{c-src}$ from crypt cell cytoskeletons is significantly higher than that in cytoskeletons from villus tip cell cytoskeletons. Moreover, nearly 70% of pp60$\sp{c-src}$ activity in crypt cells localizes to the cytoskeleton. In contrast, less than 20% of pp60$\sp{c-src}$ in villus tip cells associates with the cytoskeleton. In addition, cytoskeletal-associated pp60$\sp{c-src}$ appears to have higher specific protein tyrosine kinase activity than soluble pp60$\sp{c-src}$ in crypt cells than villus tip cells. The data suggest that active pp60$\sp{c-src}$ located on the crypt cytoskeleton may be responsible for phosphorylating proteins on tyrosine and regulating growth and differentiation of the cells.In certain cell types pp60$\sp{src}$ has been shown to act as an upstream activator of MAP kinases. Our data suggest that p41$\sp{\rm mapk}$, a 57 kD MAP kinase, and a 32 kD MBP kinase are regulated during differentiation of the intestinal epithelium. p41$\sp{\rm mapk}$ is expressed in both crypt and villus tip cells but is only active in the crypt cells where it is phosphorylated on tyrosine. The 57 kD MAP kinase and the 32 kD MBP kinase display higher MBP phosphotransferase activity in the villus tip cells. Immunofluorescence studies revealed that in addition to being localized in the nuclei of both tip and crypt cells, p41$\sp{\rm mapk}$ is also cytoplasmic and re-distributes during differentiation and colocalizes with phosphotyrosine containing proteins at the junctional complex.These results implicate cytoskeletal-associated protein tyrosine kinases including pp60$\sp{c-src}$ and MAP kinases as key regulatory molecules in normal development of intestinal epithelium.


Biology, Cell

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