Molecular Analysis of the Heterogeneity among the P-Family of Alpha-1-Antitrypsin Alleles

The rare P-family of α1-antitrypsin (α1AT) variants is defined by the position of migration of the α1AT protein on isoelectric focusing of serum (IEF) between the common M and S variants. To begin to examine the molecular heterogeneity among the P-type alleles, two unrelated subjects and their families identified by IEF to be carrying a P allele were analyzed. The first, Plowell, is a deficiency allele associated with reduced serum α1AT levels, and the second, Psaint albans is associated with normal serum levels. DNA sequence analysis of Plowell the more anodal of the two variants on IEF analysis, showed that it differed from the normal M1(Val213) allele by a single base and amino acid substitution Asp256 GAT → Val GTT. In contrast, Psaint albans, a slightly more cathodally positioned variant on IEF analysis, differed from the coding exons of the normal M1(Val213) allele by two mutations, Asp341GAC → Asn AAC, and a silent substitution in the same codon as the Piowell variant, Asp256 GAT → Asp GAC. Evaluation of Plowell mRNA transcripts by Northern and cytoblot analyses demonstrated they were of normal size and amount, and Plowell mRNA transcripts could be translated normally in vitro. Retroviral insertion of the Plowell cDNA into the genome of 3T3 fibroblasts demonstrated that it directed the synthesis of α1AT, but at levels 24% that of the Psaint albans cDNA or the normal M1 (Val213) cDNA, with a pattern of biosynthesis consistent with the concept that the Plowell α1 AT deficiency state results from intracellular degradation of the newly synthesized Plowell protein. In the context that the serum α1AT deficiency associated with other α1 AT deficiency mutations resulting from intracellular degradation of α1AT can be overcome by administering estrogenlike drugs, administration of tamoxifen to a subject with the PlowellZ phenotype resulted in α1AT serum levels rising 48% over a 5-month period, from below the threshold for protection from emphysema (11 µM) to above that threshold.