Even after action of thyrotropin (TSH) on its cell surface receptor, thyroid hormone synthesis requires the presence of multiple different molecules at the apical or basolateral plasma membranes of thyrocytes. These secretory and membrane proteins are delivered to the cell surface by intracellular transport through the secretory pathway. Successful/unsuccessful of several different proteins and cell types are proposed as models to investigate questions within this theme.
Aim 1 of this grant will focus on molecular studies of thyroglobulin (Tg), a secretory protein that has been implicated as defective for transport in certain kindreds with congenital goitrous hypothyroidism. The investigators have obtained the first stable expression of full-length non-mutant Tg in a heterologous cell type. Feasibility studies support a proposal to now produce and test site-specific mutations in the Tg backbone,, based on naturally-occurring Tg mutants, as well as mutations to identify functional domains in Tg folding and assembly.
Aim 2 of this grant will examine aspects of quality control of protein export from the endoplasmic reticulum (ER). Studies in the Progress Report demonstrate new systems being developed to compare the fates of identical secretory proteins expressed in thyrocytes versus other cell types. Feasibility studies support a plan to test their hypothesis of cell-type specific differences in the rates and efficiency of protein export and ER- associated protein degradation. The last Aim of this grant will pursue the question of targeting signals for polarized traffic of thyroid proteins expressed in the MDCK cell line, which serves as the best system available for studies of protein sorting in polarized epithelial cells.
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