Deletions within a CA-repeat-rich region of intron 4 of the human SP-B gene affect mRNA splicing.
Length variants within a CA-repeat-rich region of intron 4 of the human SP-B (pulmonary surfactant protein-B) gene are associated with several lung diseases. The hypothesis that SP-B intron 4 affects mRNA splicing was studied. SP-B minigenes containing exons 1-6 with a normal-sized intron 4 (pBi4normal) or intron 4 containing deletions (pBi4del) of 193, 211, 264 or 340 bp were expressed in CHO (Chinese hamster ovary) cells by transient transfection. Two forms of SP-B transcripts, normal and incompletely spliced, were detected. With pBi4normal, normal-sized SP-B mRNA was the predominant form and a very low amount of incompletely spliced mRNA was present, whereas with the pBi4del variants the amount of normal SP-B mRNAs was lower and the amount of incompletely spliced mRNA was relatively high. Reverse transcription-PCR results and sequencing data indicated that the incompletely spliced SP-B RNA contained intron 4 sequence, and this incompletely spliced RNA was also observed in normal lung. Lung cancer tissues with intron 4 deletions exhibited a larger amount of abnormally spliced RNAs compared with normal lung tissue or cancerous tissue with normal-sized intron 4. The results indicate that intron 4 length variants affect SP-B mRNA splicing, and that this may contribute to lung disease.
Lin
Zhenwu
Z
Department of Cellular and Molecular Physiology, The Pennsylvania State University College of Medicine, Hershey, PA 17033, USA.
Thomas
Neal J
NJ
Wang
Yunhua
Y
Guo
Xiaoxuan
X
Seifart
Carola
C
Shakoor
Hasan
H
Floros
Joanna
J
D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D002460:Cell Line; D045744:Cell Line, Tumor; D003001:Cloning, Molecular; D006224:Cricetinae; D015972:Gene Expression Regulation, Neoplastic / Q000235:genetics; D006801:Humans; D007438:Introns / Q000235:genetics*; D008168:Lung / Q000166:cytology / Q000378:metabolism; D008175:Lung Neoplasms / Q000235:genetics / Q000473:pathology; D008969:Molecular Sequence Data; D037701:Pulmonary Surfactant-Associated Protein B / Q000235:genetics* / Q000378:metabolism; D012326:RNA Splicing / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D038621:Regulatory Sequences, Ribonucleic Acid / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017384:Sequence Deletion / Q000235:genetics*
D037701:Pulmonary Surfactant-Associated Protein B; D012333:RNA, Messenger; D038621:Regulatory Sequences, Ribonucleic Acid