Molecular mechanism of growth hormone (GH) deficiency in the spontaneous dwarf rat: detection of abnormal splicing of GH messenger ribonucleic acid by the polymerase chain reaction.
The spontaneous dwarf rat (SDR) is an experimental model for pituitary dwarfism with an autosomal recessive mode of inheritance. In the pituitary of the SDR, neither GH cells nor GH protein were detected by immunological methods, but SDR pituitary has a very small amount of GH mRNA. In the present study the effects of GH-releasing factor and an analog of its second messenger, (Bu)2cAMP, on the total accumulation and secretion of GH were studied in primary cultures from SDR pituitaries. GH-releasing factor and (Bu)2cAMP increased the amount of GH mRNA, but neither accumulation nor secretion of immunoreactive GH was detected. These results suggested that the mutation is not manifest during transcription, but at a later stage. The GH gene of SDR was cloned, and its sequence was determined. A point substitution was found in the consensus sequence of the 3' splice site of the third intron. Judging from this point mutation, one would predict an abnormal splicing and a 1-base deletion in the GH mRNA. Therefore, the fragment of SDR GH cDNA in which the deletion was predicted was amplified by the polymerase chain reaction. Its sequence analysis confirmed that the SDR GH mRNA had 1 base deletion which must cause a premature translational termination of GH mRNA. These results demonstrate that GH deficiency in SDR is caused by a point mutation in the GH gene producing an abnormal splicing of GH mRNA.
Takeuchi
T
T
Department of Anatomy, Jikei University School of Medicine, Tokyo, Japan.
Suzuki
H
H
Sakurai
S
S
Nogami
H
H
Okuma
S
S
Ishikawa
H
H
D000818:Animals; D001696:Biomechanical Phenomena; D003994:Bucladesine / Q000494:pharmacology; D005576:Colforsin / Q000494:pharmacology; D004274:DNA, Recombinant; D004392:Dwarfism / Q000235:genetics* / Q000378:metabolism; D005796:Genes; D013006:Growth Hormone / Q000172:deficiency* / Q000235:genetics; D013007:Growth Hormone-Releasing Hormone / Q000494:pharmacology; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000235:genetics*; D051381:Rats; D011922:Rats, Mutant Strains / Q000235:genetics*
D004274:DNA, Recombinant; D012333:RNA, Messenger; D005576:Colforsin; D003994:Bucladesine; D013006:Growth Hormone; D013007:Growth Hormone-Releasing Hormone