FHL2 suppresses growth and differentiation of the colon cancer cell line HT-29

Abstract

Abstract. Four and a half LIM domain protein 2 (FHL2) can interact with many proteins and regulates different cellular processes, including proliferation and differentiation. FHL2 expression is often deregulated in cancer and may act as both tumor-promoter or tumor-suppressor depending on the type of cancer. Thus, a previous study found that increased FHL2 expression in colon cancer and suppression of FHL2 in a colon cancer cell line with endogenously high FHL2 expression inhibited tumor growth. We applied the opposite strategy, an FHL2 expression plasmid was stably transfected into HT-29 cells, a colon carcinoma cell line which exhibits very low basal levels of FHL2. Stable expression of FHL2 in HT-29 cells induced a G2/M arrest and inhibited anchorage-dependent and -independent growth in vitro. Further, FHL2 expressing HT-29 cell clones revealed significantly higher expression of the differentiation marker E-cadherin but reduced activity of the transcription factor NF-κB, which is known to promote colon cancer progression. These findings further underscore the complex role of FHL2 in tumorigenicity, with even different effects on cellular functions of cancer cell lines derived from the same type of tumor and distinctly suggest caution regarding therapeutic strategies targeting FHL2 to treat (colon) cancer. IntroductionColorectal cancer (CRC) is one of the most frequent human tumors and the third leading cause of cancer death (1). The prognosis of CRC, especially advanced cancer, has not substantially improved although the molecular mechanisms of CRC have become better understood in the past two decades (2).Four and a half LIM protein 2 (FHL2), also known as down-regulated in rhabdomyosarcoma LIM (DRAL) domain protein, is a member of a small family of five proteins with four and a half LIM domains (3). The abbreviation LIM represents the names of three transcription factors (Lin-11, Isl-1 and Mec-3) in which such a domain was first identified (4). The LIM domains are double zinc finger motifs that play multiple roles in protein-protein interaction (5). FHL2 can interact with >50 different proteins with diverse functions such as receptors, enzymes, transcription factors, cofactors or splicing factors (6). Thus, FHL2 is involved in the regulation of various cellular processes including proliferation, differentiation, migration and contraction (7-10). FHL2 can function as repressor or activator of transcriptional activity, and noteworthy, depending on the cell type even the expression of the same gene or distinguished cellular functions, respectively, may be regulated in different ways. Thus, FHL2 promotes expression of ·-smooth-muscle actin (·-SMA) and contractility in myofibroblasts of the skin (9), but inhibits ·-SMA expression and contractility of cardiac muscle cells (11).Also the role of FHL2 in tumorigenesis is complex, since it may act as an oncoprotein or as a tumor suppressor depending on the type of cancer (6,12). FHL2 was first identified in human rhabdomyosarcoma cells as a downregulated gene indicati...

Targeting FHL2‑E‑cadherin axis by miR‑340‑5p attenuates colon cancer cell migration and invasion

“…Analysis of GSE97023 microarray dataset revealed that commonly used colon cancer cell lines have two different levels of FHL2 expression, indicating that colon cancer might have two different levels of FHL2 expression depending on the colon cell lines. This observation is in line with Amann et al ( 37 ) reporting that HT-29 cell line has lower levels of FHL2 expression compared to SW48, CaCo, Lovo, SW480 and HCT116 colon cancer cell lines. We utilized two different cells lines (HT-29 and AZ-97) representing low and relatively high FHL2 expressing colon cancer cells for our subsequent stress related study.…”

Effects of the FHL2 gene on the development of subcutaneous and intramuscular adipocytes in goats

“…Interestingly, FHL2 increased the expression of LPL in subcutaneous adipocytes, but decreased LPL expression in intramuscular adipocytes. Unlike this study, FHL2 inhibits colon cancer cell line HT-29 cells and neuronal cell growth and differentiation [40,41]. The above results show that FHL2 has different effects on the differentiation of different adipocytes, and the results of this study show that FHL2 promotes intramuscular and subcutaneous adipocyte differentiation in goats, and the molecular mechanism of regulation is different.…”

RNA sequencing of long-term label-retaining colon cancer stem cells identifies novel regulators of quiescence

“…In addition, such strategies may also enable cell cycle activation of qCSCs in p53 mutant tumors. Differential gene expression analysis, comparing PKH26 Negative and PKH26 Positive cells, identified the negative cell cycle regulators AKAP12 ( Gelman, 2010 ; Lin et al., 2000 ; Liu et al., 2011 ; Reggi and Diviani, 2017 ), CD82 ( Hur et al, 2016 ), CDKN1A ( El-Deiry, 1993 ; Vogelstein et al., 2000 ; Wade Harper, 1993 ; Xiong et al., 1993 ), FHL2 ( Hellerbrand, 2010 ; Labalette et al, 2008 ; Lee et al., 2006 ; Martin et al., 2007 ), GPX3 ( An et al., 2018 ; Barrett et al, 2013 ; Wang et al, 2012 ), KIAA0247 ( Huang et al, 2011 ; Polato et al, 2014 ), LCN2 ( Chakraborty et al., 2012 ; Chiang et al, 2016 ; Kim et al., 2017 ), TFF2 ( Bossenmeyer-Pourié et al, 2002 ; Dubeykovskaya et al, 2016 , 2019 ; Thim, 1997 ; Tu et al, 2009 ), UNC5B ( Huang et al., 2020 ; Kong et al., 2016 ; Okazaki, 2011 ), and ZMAT3 ( Bersani et al., 2014 ; Hellborg et al, 2001 ) to be enriched in qCSCs ( Figure 4 A and Data S1 ). Significantly, each of these genes is a target of p53 ( Bersani et al., 2014 ; Fischer, 2017 ; Gelman, 2010 ; Lane, 1992 ; Lee et al., 2006 ; Liu et al., 2011 ; Marreiros et al., 2005 ; Miyamoto et al, 2016 ; Polato et al, 2014 ; Rouillard et al., 2016 ; Soutto et al, 2014 ), and with the exceptions of LCN2 and ZMAT 3, is associated with reduced survival in colorectal cancer (CRC) ( Figure 4 B).…”