Global profiling and molecular characterization of alternative splicing events misregulated in lung cancer.
Alternative splicing (AS) is a widespread mechanism underlying the generation of proteomic and regulatory complexity. However, which of the myriad of human AS events play important roles in disease is largely unknown. To identify frequently occurring AS events in lung cancer, we used AS microarray profiling and reverse transcription-PCR (RT-PCR) assays to survey patient-matched normal and adenocarcinoma tumor tissues from the lungs of 29 individuals diagnosed with non-small cell lung cancer (NSCLC). Of 5,183 profiled alternative exons, four displayed tumor-associated changes in the majority of the patients. These events affected transcripts from the VEGFA, MACF1, APP, and NUMB genes. Similar AS changes were detected in NUMB and APP transcripts in primary breast and colon tumors. Tumor-associated increases in NUMB exon 9 inclusion correlated with reduced levels of NUMB protein expression and activation of the Notch signaling pathway, an event that has been linked to tumorigenesis. Moreover, short hairpin RNA (shRNA) knockdown of NUMB followed by isoform-specific rescue revealed that expression of the exon 9-skipped (nontumor) isoform represses Notch target gene activation whereas expression of the exon 9-included (tumor) isoform lacks this activity and is capable of promoting cell proliferation. The results thus reveal widespread AS changes in NSCLC that impact cell signaling in a manner that likely contributes to tumorigenesis.
Banting and Best Department of Medical Research, University of Toronto, Donnelly Centre, 160 College Street, Toronto, Ontario, Canada M5S 3E1.
Christine M
CM
Misquitta-Ali
Edith
E
Cheng
Dave
D
O'Hanlon
Ni
N
Liu
C Jane
CJ
McGlade
Ming Sound
MS
Tsao
Benjamin J
BJ
Blencowe
D000230:Adenocarcinoma / Q000235:genetics; D017398:Alternative Splicing; D016564:Amyloid beta-Protein Precursor / Q000235:genetics; D001483:Base Sequence; D001943:Breast Neoplasms / Q000235:genetics / Q000378:metabolism; D002289:Carcinoma, Non-Small-Cell Lung / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D016022:Case-Control Studies; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D003110:Colonic Neoplasms / Q000235:genetics / Q000378:metabolism; D005091:Exons; D005260:Female; D020869:Gene Expression Profiling; D055785:Gene Knockdown Techniques; D006801:Humans; D008175:Lung Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D008297:Male; D008565:Membrane Proteins / Q000037:antagonists & inhibitors / Q000235:genetics; D008840:Microfilament Proteins / Q000235:genetics; D009419:Nerve Tissue Proteins / Q000037:antagonists & inhibitors / Q000235:genetics; D020411:Oligonucleotide Array Sequence Analysis; D009928:Organ Specificity; D011247:Pregnancy; D012333:RNA, Messenger / Q000235:genetics; D012334:RNA, Neoplasm / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics; D051880:Receptors, Notch / Q000378:metabolism; D015398:Signal Transduction; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics
C545487:APP protein, human; D016564:Amyloid beta-Protein Precursor; C095003:MACF1 protein, human; D008565:Membrane Proteins; D008840:Microfilament Proteins; D009419:Nerve Tissue Proteins; C113026:NUMB protein, human; D012333:RNA, Messenger; D012334:RNA, Neoplasm; D034741:RNA, Small Interfering; D051880:Receptors, Notch; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A