Inducible intestine-specific deletion of Krüppel-like factor 5 is characterized by a regenerative response in adult mouse colon.
Krüppel-like factor 5 (KLF5) is a pro-proliferative transcriptional regulator primarily expressed in the intestinal crypt epithelial cells. Constitutive intestine-specific deletion of Klf5 is neonatal lethal suggesting a crucial role for KLF5 in intestinal development and homeostasis. We have previously shown Klf5 to play an active role regulating intestinal tumorigenesis. Here we examine the effect of inducible intestine-specific deletion of Klf5 in adult mice. Klf5 is lost from the intestine beginning at day 3 after the start of a 5-day treatment with the inducer tamoxifen. Although the mice have no significant weight loss or lethality, the colonic tissue shows signs of epithelial distress starting at day 3 following induction. Accompanying the morphological changes is a significant loss of proliferative crypt epithelial cells as revealed by BrdU or Ki67 staining at days 3 and 5 after start of tamoxifen. We also observed a loss of goblet cells from the colon and Paneth cells from the small intestine upon induced deletion of Klf5. In addition, loss of Klf5 from the colonic epithelium is accompanied by a regenerative response that coincides with an expansion in the zone of Sox9 expression along the crypt axis. At day 11, both proliferation and Sox9 expression return to baseline levels. Microarray and quantitative PCR analyses reveal an up-regulation of several regeneration-associated genes (Reg1A, Reg3G and Reg3B) and down-regulation of many Klf5 targets (Ki-67, cyclin B, Cdc2 and cyclin D1). Sox9 and Reg1A protein levels are also increased upon Klf5 loss. Lentiviral-mediated knockdown of KLF5 and exogenous expression of KLF5 in colorectal cancer cell lines confirm that Sox9 expression is negatively regulated by KLF5. Furthermore, ChIP assays reveal a direct association of KLF5 with both the Sox9 and Reg1A promoters. We have shown that disruption of epithelial homeostasis due to Klf5 loss from the adult colon is followed by a regenerative response led by Sox9 and the Reg family of proteins. Our study demonstrates that adult mouse colonic tissue undergoes acute physiological changes to accommodate the loss of Klf5 withstanding epithelial damage further signifying importance of Klf5 in colonic homeostasis.
Department of Medicine, Stony Brook University School of Medicine, HSC-T16 Room 020, Stony Brook, NY 11794, USA. Electronic address: Mandayam.Nandan@stonybrookmedicine.edu.
Mandayam O
MO
Nandan
Department of Medicine, Stony Brook University School of Medicine, HSC-T16 Room 020, Stony Brook, NY 11794, USA. Electronic address: Amr.Ghaleb@stonybrookmedicine.edu.
Amr M
AM
Ghaleb
Department of Medicine, Stony Brook University School of Medicine, HSC-T16 Room 020, Stony Brook, NY 11794, USA. Electronic address: yangliu5@ic.sunysb.edu.
Yang
Y
Liu
Department of Medicine, Stony Brook University School of Medicine, HSC-T16 Room 020, Stony Brook, NY 11794, USA. Electronic address: Agnieszka.Bialkowska@stonybrookmedicine.edu.
Agnieszka B
AB
Bialkowska
Division of Digestive Diseases, Department of Medicine, Emory University School of Medicine, 615 Michael Street, Atlanta, GA 30322, USA. Electronic address: Beth.McConnell@data2impact.com.
Beth B
BB
McConnell
Department of Pathology, Stony Brook University School of Medicine, BST-9, Stony Brook Medicine, Stony Brook, NY 11794-8691, USA. Electronic address: Kenneth.Shroyer@stonybrookmedicine.edu.
Kenneth R
KR
Shroyer
Department of Morphogenesis and Intracellular Signalling, Institut Curie-CNRS, Paris, France. Electronic address: Sylvie.Robine@curie.fr.
Sylvie
S
Robine
Department of Medicine, Stony Brook University School of Medicine, HSC-T16 Room 020, Stony Brook, NY 11794, USA. Electronic address: Vincent.Yang@stonybrookmedicine.edu.
Vincent W
VW
Yang
D000818:Animals; D018931:Antineoplastic Agents, Hormonal / Q000494:pharmacology; D016203:CDC2 Protein Kinase / Q000378:metabolism; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D003106:Colon / Q000502:physiology*; D015179:Colorectal Neoplasms / Q000378:metabolism; D019926:Cyclin B / Q000378:metabolism; D019938:Cyclin D1 / Q000378:metabolism; D015536:Down-Regulation; D020397:Goblet Cells / Q000187:drug effects; D045325:HCT116 Cells; D057809:HEK293 Cells; D006801:Humans; D019394:Ki-67 Antigen / Q000378:metabolism; D051741:Kruppel-Like Transcription Factors / Q000235:genetics* / Q000378:metabolism; D051847:Lithostathine / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D000073718:Pancreatitis-Associated Proteins; D019879:Paneth Cells / Q000187:drug effects; D011401:Promoter Regions, Genetic; D011506:Proteins / Q000378:metabolism; D034622:RNA Interference; D034741:RNA, Small Interfering; D012038:Regeneration / Q000235:genetics*; D055758:SOX9 Transcription Factor / Q000378:metabolism; D017384:Sequence Deletion; D015398:Signal Transduction / Q000235:genetics; D013629:Tamoxifen / Q000494:pharmacology; D015854:Up-Regulation
D018931:Antineoplastic Agents, Hormonal; D019926:Cyclin B; D019394:Ki-67 Antigen; C118391:Klf5 protein, mouse; D051741:Kruppel-Like Transcription Factors; D051847:Lithostathine; D000073718:Pancreatitis-Associated Proteins; D011506:Proteins; C497617:REG1A protein, human; C494437:REG3G protein, human; D034741:RNA, Small Interfering; C490253:Reg3b protein, mouse; D055758:SOX9 Transcription Factor; C527933:Sox9 protein, mouse; D013629:Tamoxifen; D019938:Cyclin D1; D016203:CDC2 Protein Kinase
Colon; Cre recombinase; Differentiation; Epithelium; Inducible deletion; Intestine; KLF5; Proliferation; Reg1A; Regeneration; Sox9