JCB_201708064 1335..1352

Four-and-a-half LIM (FHL) protein 1 (FHL1) belongs to the FHL protein family, which consists of four members, FHL1, FHL2, FHL3, and FHL5 in humans. All these proteins are characterized by the tandem arrangement of four and a half highly conserved LIM domains. LIM domains mediate protein–protein interactions and are involved in linking proteins with both the actin cytoskeleton and the transcriptional machinery (Kadrmas and Beckerle, 2004; Shathasivam et al., 2010). FHL1 is highly expressed in skeletal muscle and heart (Greene et al., 1999) and has been associated with skeletal muscle myopathies and several cardiovascular diseases (Cowling et al., 2008; Willis et al., 2016). Interestingly, FHL1 is markedly down-regulated in a variety of cancers including lung (Niu et al., 2012), liver (Ding et al., 2009), breast (Ding et al., 2011), colon, renal (Li et al., 2008), and gastric cancers (Xu et al., 2012). FHL1 was previously identified as a tumor suppressor protein, which acts to inhibit tumor cell growth and migration. Recently, our study (Xu et al., 2017) showed that FHL1 leads to radiation resistance in cancer cells by inhibiting CDC25C activity. Moreover, increased expression of FHL1 led to significantly poorer disease-free survival and overall survival rates for breast cancer patients who received radiotherapy, indicating that the role and mechanism of FHL1 in cancer progression is more complex and diverse than was previously thought. Whether FHL1 is an implicit tumor cell growth suppressor needs to be questioned and investigated. Additionally, although it is certain that FHL1 expression is down-regulated in many cancers, the posttranslational modification of FHL1 and the potential role of such modifications in cancer progression remain unclear. Previous research has indicated that FHL1 localizes to the nucleus and focal adhesions via integrin activation, where it then functions to promote cell spreading and migration (Robinson et al., 2003). Upon activation, integrins subsequently activate cytoplasmic kinases and cytoskeletal signaling cascades including enzymes (e.g., focal adhesion kinase [FAK], Src, and Rho GTPases) and adapters (e.g., paxillin; Guo and Giancotti, 2004; Harburger and Calderwood, 2009). With respect to FHL1, the components of the integrin-dependent signaling pathways that are responsible for FHL1 localization to the nucleus and focal adhesions and the functions of FHL1 at these specific locations remain unclear. Kindlin-2, a member of the kindlin protein family, is considered as an essential regulator of integrin activation and integrin-mediated cell–ECM adhesion (Larjava et al., 2008; Ma et al., 2008). Kindlin-2 is reported to act as an adapter protein, and as an important member of focal adhesion proteins, it interacts with and recruits migfilin (a LIM-containing protein) to cell–matrix adhesions and participates in the orchestration of actin assembly. Thus, we hypothesize that FHL1 is recruited to focal adhesions by interacting with kindlin-2. FHL1 has been recognized for a long time as a tumor suppressor protein that associates with both the actin cytoskeleton and the transcriptional machinery. We present in this study a paradigm that phosphorylated FHL1 functions as an oncogenic protein by promoting tumor cell proliferation. The cytosolic tyrosine kinase Src interacts with and phosphorylates FHL1 at Y149 and Y272, which switches FHL1 from a tumor suppressor to a cell growth accelerator. Phosphorylated FHL1 translocates into the nucleus, where it binds to the transcription factor BCL AF1 and promotes tumor cell growth. Importantly, the phosphorylation of FHL1 is increased in tissues from lung adenocarcinoma patients despite the downregulation of total FHL1 expression. Kindlin-2 was found to interact with FHL1 and recruit FHL1 to focal adhesions. Kindlin-2 competes with Src for binding to FHL1 and suppresses Src-mediated FHL1 phosphorylation. Collectively, we demonstrate that FHL1 can either suppress or promote tumor cell growth depending on the status of the sites for phosphorylation by Src. Src-mediated phosphorylation converts FHL1 from tumor suppressor to tumor promoter