Quantitative expression patterns of multidrug-resistance P-glycoprotein (MDR1) and differentially spliced cystic-fibrosis transmembrane-conductance regulator mRNA transcripts in human epithelia.
P-glycoprotein (MDR1), that confers multidrug resistance in cancer, and the cystic-fibrosis transmembrane-conductance regulator (CFTR), that is causative defective in cystic fibrosis, belong to the family of ATP-binding transport proteins. The expression of MDR1 and CFTR in human epithelial tissues and the cell lines T84 and HT29 was estimated by primer-directed reverse transcription (RT) and subsequent monitoring of the kinetics of cDNA product formation during the polymerase chain reaction (PCR). MDR1 mRNA was found in high levels, 15-50 amol mRNA/microgram RNA, in the intestine, kidney, liver and placenta, and in low levels, 0.2 amol/microgram RNA, in respiratory epithelium. Large amounts of CFTR mRNA were measured in the gastrointestinal tract, whereas the kidney, as the phenotypically normal organ, and the lung, as the most severely affected organ in cystic fibrosis, both contained low amounts, 3 amol CFTR/microgram RNA. CFTR transcript levels of 1-5 amol/microgram RNA were determined in lymphocytes and lymphoblast cell lines, suggesting that lymphoblasts are an accessible source for the study of the molecular pathogenesis of cystic fibrosis. When transcripts were scanned by overlapping RT/PCR analyses, only transcript of expected size was detected for MDR1 mRNA, where variable in-frame deletions of either exon 4, 9 or 12 were observed in CFTR mRNA. The complete loss of single exons was seen at proportions of 1-40% in all investigated tissues and cell lines with large donor-to-donor variation. Exons 9 and 12 of the CFTR gene encode parts of the evolutionarily well-conserved first nucleotide-binding fold including the two Walker motifs. Alternative splicing may give rise to various CFTR forms of different function and localization.
Bremer
S
S
Abteilung Biophysikalische Chemie, Medizinische Hochschule Hannover, Federal Republic of Germany.
Hoof
T
T
Wilke
M
M
Busche
R
R
Scholte
B
B
Riordan
J R
JR
Maass
G
G
Tümmler
B
B
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D001483:Base Sequence; D018288:Carcinoma, Small Cell / Q000235:genetics; D002478:Cells, Cultured; D002872:Chromosome Deletion; D003550:Cystic Fibrosis / Q000235:genetics*; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D004247:DNA / Q000235:genetics*; D004273:DNA, Neoplasm / Q000235:genetics / Q000302:isolation & purification; D004351:Drug Resistance / Q000235:genetics*; D004848:Epithelium / Q000502:physiology; D005091:Exons; D005634:Fructose-Bisphosphate Aldolase / Q000235:genetics; D006098:Granulocytes / Q000502:physiology; D006801:Humans; D007700:Kinetics; D007963:Leukocytes, Mononuclear / Q000502:physiology; D008175:Lung Neoplasms / Q000235:genetics; D008562:Membrane Glycoproteins / Q000235:genetics*; D008565:Membrane Proteins / Q000235:genetics*; D008969:Molecular Sequence Data; D009298:Nasal Polyps / Q000503:physiopathology; D009838:Oligodeoxyribonucleotides; D016133:Polymerase Chain Reaction / Q000379:methods; D012326:RNA Splicing; D012333:RNA, Messenger / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; C505032:CFTR protein, human; D004273:DNA, Neoplasm; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; D009838:Oligodeoxyribonucleotides; D012333:RNA, Messenger; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D004247:DNA; D005634:Fructose-Bisphosphate Aldolase