Inefficient membrane targeting, translocation, and proteolytic processing by signal peptidase of a mutant preproparathyroid hormone protein.
A preproparathyroid hormone allele from a patient with familial isolated hypoparathyroidism was shown to have a single point mutation in the hydrophobic core of the signal sequence. This mutation, changing a cysteine to an arginine codon at the -8 position of the signal peptide, was associated with deleterious effects on the processing of preproparathyroid hormone to proparathyroid hormone in vitro. To examine the biochemical consequence(s) of this mutation, proteins produced by cell-free translation of wild-type and mutant cRNAs were used in assays that reconstitute the early steps of the secretory pathway. We find that the mutation impairs interaction of the nascent protein with signal recognition particle and the translocation machinery. Moreover, cleavage of the mutant signal sequence by solubilized signal peptidase is ineffective. The consequence of this mutation on processing and secretion of parathyroid hormone is confirmed in intact cells by pulse-chase experiments following transient expression of the mutant protein in COS-7 cells. The inability of the mutant signal sequence, however, to interfere with the targeting and processing of other secreted proteins does not support obstruction of the translocation apparatus as the mechanism underlying the dominant mode of inheritance of hypoparathyroidism in this family.