Differential processing of hormone precursor. Independent production of somatostatins 14 and 28 in transfected neuroblastoma 2A cells.
Neuro 2A cells infected with a retroviral vector carrying human prosomatostatin cDNA expressed and processed correctly the precursor into somatostatins-14 and -28 [(1989) EMBO J. 8, 2911-2916]. In order to study the mechanisms by which the active hormone sequences arise, site directed mutagenesis was performed on either the dibasic (ArgLys) or monobasic (Arg) cleavage sites involved in the production of somatostatins-14 and -28, respectively. Radioimmunochemical analysis of the somatostatin-related products indicated that replacement of either Arg-2-Lys-1 by Asn-2-Asn-1 or of Arg-15 by Asn-15 resulted in the exclusive production of either somatostatin-28 or -14, respectively. Moreover only prosomatostatin[1-76] was detected and no somatostatin-28[1-12] could be measured in cell extracts. Selective suppression of either somatostatin-14 or somatostatin-28 release by mutation did not affect the level of production of the other hormone but resulted in a correlative increase of unprocessed prosomatostatin. It is concluded that in this cell type (i) somatostatin-14 is exclusively generated by dibasic cleavage at the Arg-2-Lys-1 site of the intact precursor with concomitant production of prosomatostatin[1-76], and (ii) no direct interactions between the monobasic and dibasic processing domains occur.
Brakch
N
N
Groupe de Neurobiochimie Cellulaire et Moléculaire, Université Pierre et Marie Curie, Paris, France.
Rholam
M
M
Nault
C
C
Boileau
G
G
Cohen
P
P
D000595:Amino Acid Sequence; D002851:Chromatography, High Pressure Liquid; D004252:DNA Mutational Analysis; D066298:In Vitro Techniques; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011498:Protein Precursors / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D011994:Recombinant Proteins; D013004:Somatostatin / Q000378:metabolism*; D014162:Transfection
D011498:Protein Precursors; D011994:Recombinant Proteins; D013004:Somatostatin; C028456:prosomatostatin