Mammalian Krüppel-like factors in health and diseases

Abstract

The Krüppel-like factor (KLF) family of transcription factors regulates diverse biological processes that include proliferation, differentiation, growth, development, survival, and responses to external stress. Seventeen mammalian KLFs have been identified, and numerous studies have been published that describe their basic biology and contribution to human diseases. KLF proteins have received much attention because of their involvement in the development and homeostasis of numerous organ systems. KLFs are critical regulators of physiological systems that include the cardiovascular, digestive, respiratory, hematological, and immune systems and are involved in disorders such as obesity, cardiovascular disease, cancer, and inflammatory conditions. Furthermore, KLFs play an important role in reprogramming somatic cells into induced pluripotent stem (iPS) cells and maintaining the pluripotent state of embryonic stem cells. As research on KLF proteins progresses, additional KLF functions and associations with disease are likely to be discovered. Here, we review the current knowledge of KLF proteins and describe common attributes of their biochemical and physiological functions and their pathophysiological roles.

Figure 1. Phylogenetic tree of human KLFs

Multiple sequence alignment and phylogenetic analysis were performed using the ClustalW tool, Version 2.0.12. Analysis was conducted on full-length protein sequences of the 17 human KLF proteins. Structural analysis corresponded with the division of KLFs into distinct groups that have functional similarities.

Figure 2. Protein structure of human KLF family members

KLF proteins are grouped according to common structural and functional domains. KLFs are highly homologous in their carboxyl-terminal DNA-binding regions, which contain three C₂H₂ zinc finger motifs. The family members were grouped based on: (1) the ability to bind acetylases (KLFs 1, 2, 4, 5, 6, and 7); (2) the presence of a CtBP-binding site (KLFs 3, 8, and 12); or (3) the presence of a Sin3A-binding site (KLFs 9, 10, 11, 13, 14, and 16). Established sites of acetylation are marked by stars.

Figure 3. Localization of Klf4 and Klf5 in the mouse colon

Immunofluorescence staining of Klf 4 or 5 (green) with the proliferation marker, Ki67 (red), was conducted on mouse colon. Klf4 (green) is present in the nuclei of terminally differentiated epithelial cells in the upper regions of colonic crypts. In contrast, Klf5 is localized to nuclei of proliferating epithelial cells at the base of the crypts. Ki67 highlights regions of active proliferation. Although Klf4 and Ki67 staining patterns do not overlap, Klf5 and Ki67 exhibit considerable co-localization, indicated by yellow staining.

Figure 4. Reduced mucosal hyperplasia in Klf5^+/− mice following infection with the enteric pathogen, Citrobacter rodentium

In these experiments, C57BL/6 (WT) and Klf5^+/− mice were infected with the rodent-specific pathogen, Citrobacter rodentium. Colonic tissues from uninfected and C. rodentium-infected mice were isolated at 14 days post-infection (p.i.) and were subjected to histological staining with hematoxylin and eosin (H&E) or immunofluorescence staining for expression of Klf5 (green) or the proliferation marker, Ki67 (red). Hyperplasia induced by infection was significantly reduced in the Klf5^+/− mice [from McConnell, et al. (273)].

Figure 5. Contribution of KLFs in regulating adipocyte differentiation

Pre-adipocytes progress through an early stage of differentiation in which they become committed to their fate, followed by a late stage of terminal differentiation into mature adipocytes. Various members of the KLF family contribute to transcriptional control of this process and have positive and negative effects on adipogenesis [Adapted from Nagai, Friedman and Kasuga, Editors (294)].

Figure 6. A model for the role of KLF4 in tumor suppression and oncogenesis

Following DNA damage, expression of KLF4 is activated in a p53-dependent manner. Increased levels of KLF4 lead to increased expression of p21^Cip1/Waf1 and decreased expression of BAX, with the net effect being to tip the balance away from apoptosis towards cell cycle arrest. However, in the presence of activated RAS or E1A, the growth inhibitory effects of p21^Cip1/Waf1 are negated by high levels of cyclin D1. The combination of high levels of KLF4 expression in the presence of oncogenic RAS or E1A lead to active proliferation and suppressed apoptosis, which contribute to carcinogenesis. KLF6 likewise acts as a tumor suppressor through induction of p21^Cip1/Waf1. The growth-suppressing activity of KLF6 can be offset by oncogenic RAS which induces alternative splicing of KLF6, generating a mutant form that promotes cell proliferation and suppresses apoptosis [Adapted from Ghaleb, et al. (149)].

Figure 7. Klf4 as part of a transcriptional regulatory circuit for somatic cell reprogramming

A model of the transcriptional hierarchy of the reprogramming transcription factors: Klf4, Sox2, Oct4, c-Myc. Klf4 is shown as an upstream regulator of feed-forward transcription loops—it binds the promoters of Oct4, Sox2, c-Myc and the downstream target Nanog. Klfs 2 and 5 can substitute for Klf4 in somatic cell reprogramming [Adapted from Kim, et al. (217)].