Serum alpha 1-antitrypsin deficiency associated with the common S-type (Glu264----Val) mutation results from intracellular degradation of alpha 1-antitrypsin prior to secretion.
The S-type alpha 1-antitrypsin (alpha 1AT) deficiency allele differs from the normal M1(Val213) allele by a single amino acid substitution (Glu264----Val). To evaluate the molecular pathophysiology responsible for the reduced serum levels of alpha 1AT associated with the S-type allele, alpha 1AT gene expression was examined in blood monocytes, cells which normally produce alpha 1AT, as well as murine fibroblasts modified by retroviral gene transfer to express the S-type and normal M-type human alpha 1AT genes. Northern analysis and S1 protection analysis demonstrated that monocytes of M and S homozygotes both express 1.8-kilobase alpha 1AT mRNA transcripts in comparable levels and similar in structure. Pulse-chase labeling studies demonstrated that both M and S monocytes synthesized and secreted a 52-kDa protein, but the S monocytes secreted significantly less. The cellular lysates of both M and S monocytes contained a newly synthesized 50-kDa precursor form of alpha 1AT, but the S monocytes contained reduced amounts. Pulse-chase labeling in the presence of tunicamycin, an inhibitor of core oligosaccharide addition, demonstrated that S monocytes exhibited a selective inhibition of secretion of 45-kDa nonglycosylated alpha 1AT not observed in M monocytes. Consistent with these observations, murine fibroblasts modified by retroviral gene transfer to contain an integrated human S-type alpha 1AT cDNA demonstrated reduced secretion of alpha 1AT compared with fibroblasts containing an integrated human M-type alpha 1AT cDNA and also reproduced the abnormality of alpha 1AT biosynthesis observed with S-type monocytes. Furthermore, in the presence of leupeptin, an inhibitor of cellular proteinases, the S-type modified fibroblasts demonstrated a selective augmentation of human alpha 1AT secretion not observed for the M-type. Together, these observations are consistent with the concept that the single A----T mutation of the S-type alpha 1AT gene results in reduced cellular secretion of alpha 1AT because the newly synthesized S-type alpha 1AT protein is degraded intracellularly prior to secretion.
Curiel
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DT
Pulmonary Branch, National Heart, Lung, and Blood Institute, Bethesda, Maryland 20892.
Chytil
A
A
Courtney
M
M
Crystal
R G
RG
D000483:Alleles; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D005796:Genes; D005971:Glutamates; D018698:Glutamic Acid; D006720:Homozygote; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D009000:Monocytes / Q000378:metabolism; D009154:Mutation; D010957:Plasmids; D012333:RNA, Messenger / Q000235:genetics; D014158:Transcription, Genetic; D014162:Transfection; D014633:Valine; D000515:alpha 1-Antitrypsin / Q000235:genetics* / Q000378:metabolism; D019896:alpha 1-Antitrypsin Deficiency
D005971:Glutamates; D012333:RNA, Messenger; D000515:alpha 1-Antitrypsin; D018698:Glutamic Acid; D014633:Valine