ALPHA-1-ANTITRYPSIN GENE MUTATION HOT-SPOT ASSOCIATED WITH THE FORMATION OF A RETAINED AND DEGRADED NULL VARIANT

Null alpha 1-antitrypsin (alpha 1AT) alleles represent the end of a co ntinuum of variants associated with profound alpha 1AT deficiency and an increased risk of emphysema. This study characterizes the molecular basis of QOclayton, a new example of an alpha 1AT null allele arising from a mutational hot spot in the alpha 1AT gene. The QOclayton allel e is identical to the normal M1(V213) alpha 1AT allele except for an i nsertion of a cytosine. This insertion occurs in the alpha 1AT sequenc e which normally has seven cytosines corresponding to amino acid resid ues 360 to 362. The QOclayton mutation is located in the same reiterat ed DNA sequence as the alpha 1AT QObolton deletion mutation and the in sertion mutation allele QOsaarbruecken. The QOclayton cytosine inserti on causes a 3' frameshift and results in the formation of a terminatio n codon at residue 376, the same consequence as the alpha 1AT QOmattaw a mutation (L353 T-insertion with a 3' frameshift). To determine the m olecular mechanisms responsible for the absence of alpha 1AT associate d with the QOclayton gene, an in vitro model of QOclayton was establis hed using Chinese hamster ovary cells (CHO) transfected with the QOcla yton gene. These cells were evaluated for alpha 1AT mRNA expression, p rotein synthesis and secretion. Although the QOclayton gene expresses a similar amount of alpha 1AT mRNA as compared with the normal alpha 1 AT gene, no QOclayton protein is secreted. Protein trafficking and dou ble-label immunofluorescence demonstrate that the QOclayton protein is retained in the rough endoplasmic reticulum or pre-Golgi compartment and is degraded (t(1/2) = 6.5 h). Since QOmattawa, QObolton, and QOsaa rbruecken have similar termination sites in the alpha 1AT mRNA, they m ay share a similar intracellular fate.