The dominant view in protein science is that a three-dimensional (3-D) structure is a prerequisite for protein function. In contrast to this dominant view, there are many counterexample proteins that fail to fold into a 3-D structure, or that have local regions that fail to fold, and yet carry out function. Protein without fixed 3-D structure is called intrinsically disordered. Motivated by anecdotal accounts of higher rates of sequence evolution in disordered protein than in ordered protein we are exploring the molecular evolution of disordered proteins. To test whether disordered protein evolves more rapidly than ordered protein, pairwise genetic distances were compared between the ordered and the disordered regions of 26 protein families having at least one member with a structurally characterized region of disorder of 30 or more consecutive residues. For five families, there were no significant differences in pairwise genetic distances between ordered and disordered sequences. The disordered region evolved significantly more rapidly than the ordered region for 19 of the 26 families. The functions of these disordered regions are diverse, including binding sites for protein, DNA, or RNA and also including flexible linkers. The functions of some of these regions are unknown. The disordered regions evolved significantly more slowly than the ordered regions for the two remaining families. The functions of these more slowly evolving disordered regions include sites for DNA binding. More work is needed to understand the underlying causes of the variability in the evolutionary rates of intrinsically ordered and disordered protein.

Actin filament formation and turnover within the treadmilling actin filament array at the leading edge of migrating cells are interdependent and coupled, but the mechanisms coordinating these two activities are not understood. We report that Coronin 1B interacts simultaneously with Arp2/3 complex and Slingshot (SSH1L) phosphatase, two regulators of actin filament formation and turnover, respectively. Coronin 1B inhibits filament nucleation by Arp2/3 complex and this inhibition is attenuated by phosphorylation of Coronin 1B at Serine 2, a site targeted by SSH1L. Coronin 1B also directs SSH1L to lamellipodia where SSH1L likely regulates Cofilin activity via dephosphorylation. Accordingly, depleting Coronin 1B increases phospho-Cofilin levels, and alters lamellipodial dynamics and actin filament architecture at the leading edge. We conclude that Coronin 1B's coordination of filament formation by Arp2/3 complex and filament turnover by Cofilin is required for effective lamellipodial protrusion and cell migration.

Androgen receptor (AR) activity is required for prostate cancer development and progression. Thus, there is a major impetus to understand the regulation of AR action. We and others have previously shown that AR transactivation potential is dependent on the presence of an active SWI/SNF chromatin remodeling complex. However, the mechanisms underlying SWI/SNF regulation of the AR remained unsolved. We show here that the BAF57 subunit, an accessory component of the remodeling complex, is a critical regulator of AR function. We show that BAF57 is expressed in the luminal epithelia of the prostate and is required for AR-dependent transactivation in prostatic adenocarcinoma cells. Our data reveal that BAF57 can directly bind to the AR and is recruited to endogenous AR targets upon ligand activation. Loss of BAF57 or inhibition of BAF57 function severely compromised AR activity, as observed with both exogenous and endogenous AR targets. Rescue of BAF57 function restored AR activity, thus demonstrating a specific requirement of BAF57 for AR activity. This action of BAF57 proved to be dependent on SWI/SNF ATPase function. BAF57 has previously been implicated in nuclear receptor coactivator function, and we show that, although BAF57 facilitated coactivator activity, only a selected subset required BAF57 for coactivator function. Lastly, we demonstrate that both BAF57 and BRM are required for the proliferation of AR-dependent prostatic adenocarcinoma cells. In summary, these findings identify BAF57 as a critical modulator of the AR that is capable of altering AR activity, coactivator function, and AR-dependent proliferation.

Adenomatous polyposis coli (APC) controls the direction in which cells extrude from epithelia. APC acts in the dying cell to control where microtubules target actomyosin contraction in neighboring cells that squeeze out the dying cell. APC mutations that frequently occur in colon cancer cause cells to extrude aberrantly beneath epithelia, which could enable tumor cell invasion.

The androgen receptor (AR) is a ligand-dependent transcription factor whose activity is required for prostate cancer proliferation. Because ablation of AR activity is a critical goal of prostate cancer therapy, much emphasis has been placed on understanding the accessory proteins that regulate AR function in the prostate. Several co-activators have been shown to be required for full AR activity, including histone acetyltransferases and TRAP/mediator complexes. SWI/SNF comprises a family of large, multisubunit complexes present in the cell, which contain one of two core ATPases required for nucleosome re-positioning, BRG1 or hBRM. We investigated the specific requirement of the SWI/SNF core ATPases for AR function. Using cells deficient in both BRG1 and hBRM, we show that activation of one AR target promoter, prostate-specific antigen (PSA), requires SWI/SNF chromatin remodeling for activity. A second AR target promoter, probasin, maintained a low level of activation in the absence of SWI/SNF. AR stimulation on the probasin core promoter could be partially induced with BRG1, but hBRM strongly stimulated AR activity. The PSA promoter was only induced by the restoration of hBRM. In contrast, ligand-dependent activation of the estrogen receptor was equally stimulated by BRG1 or hBRM. We demonstrate that the addition of a known enhancer region to the core PSA promoter bypasses the requirement for SWI/SNF on the PSA promoter, indicating that elements upstream of specific proximal promoters can impact the influence of the SWI/SNF complex on target gene activation. Addition of the enhancer to the probasin core promoter failed to impact the SWI/SNF requirement. In summary, SWI/SNF function potently regulates core AR target gene promoter activation, with a preference for hBRM-containing complexes. These studies highlight a role for the enhancer in altering the impact of SWI/SNF action and suggest a disparity in AR target genes for SWI/SNF requirement.

Precise control of cell-matrix adhesion is necessary for cell migration to occur. Fibroblasts lacking paxillin (PXN) or focal adhesion kinase (FAK), two core focal-adhesion components, display aberrant adhesion and decreased cell motility in vitro. Embryos deficient in genes that encode these proteins also lack proper mesoderm formation leading to embryonic lethality (Ilic et al., 1995;Hagel et al., 2002). In many types of cancer cell, focal adhesion proteins display either modulated protein expression or inappropriate regulation. For example, FAK is overexpressed in many types of cancer cells and is responsible for hyperphosphorylation of other focal-adhesion proteins (Mitra and Schlaepfer, 2006). Conversely, targeted disruption of the FAK gene in breast cancer models show that FAK is required for carcinoma formation and metastasis (Lahlou et al., 2007). Other focal-adhesion proteins have altered expression profiles in cancer models as well. Recent data suggest that the EGF-induced switch from expression of tensin-3 to the expression of the anti-adhesive molecule cten leads to increased metastasis of mammary tumor cells (Katz et al., 2007). These observations indicate that proper focal-adhesion dynamics are crucial for morphogenesis and play a significant role in cancer progression.Coronins are highly conserved F-actin binding proteins that are important for cell motility and actin dynamics (de Hostos et al., 1993;Cai et al., 2007;Foger et al., 2006). Mammalian genomes contain at least six coronin genes that can be separated into three types: type I (coronin 1A, 1B and 1C), type II (coronin 2A and 2B), and type III (coronin 7, also known as POD) (Uetrecht and Bear, 2006). Each of the coronins displays different tissue expression patterns with at least one type I coronin and POD expressed in all tissues and cell types. Type II coronins show a more restricted expression pattern, with strong enrichment in tissues containing epithelial and neuronal cell types (Cai et al., 2005).The subcellular localization of each of the coronin types is also quite distinct. Type I coronins localize primarily to the lamellipodia and some vesicular structures (Cai et al., 2005;Rosentreter et al., 2007), type II coronins localize to stress fibers and focal adhesions (Nakamura et al., 1999), and POD localizes to the Golgi apparatus (Rybakin et al., 2004). Type I coronins have a clear role in regulating cell motility, whereas the function of type II coronins remains unknown. Type I coronins such as coronin 1B coordinately regulate Arp2/3 and cofilin activities in lamellipodia (Cai et al., 2007). Coronin 1B targets Arp2/3-containing branches and replaces Arp2/3 at branches, leading to network remodeling and disassembly (Cai et al., 2008). In addition, coronin 1B is required for proper targeting of Slingshot-1L, an activating phosphatase for the ADF/cofilin family of actin-binding proteins to the rear of lamellipodia (Cai et al., 2007). It is unclear whether type II coronins execute similar functions at other cellular locations.ADF/cofil...

Persistent cellular migration requires efficient protrusion of the front of the cell, the leading edge where the actin cytoskeleton and cell-substrate adhesions undergo constant rearrangement. Rho family GTPases are essential regulators of the actin cytoskeleton and cell adhesion dynamics. Here, we examined the role of the RhoGEF TEM4, an activator of Rho family GTPases, in regulating cellular migration of endothelial cells. We found that TEM4 promotes the persistence of cellular migration by regulating the architecture of actin stress fibers and cell-substrate adhesions in protruding membranes. Furthermore, we determined that TEM4 regulates cellular migration by signaling to RhoC as suppression of RhoC expression recapitulated the loss-of-TEM4 phenotypes, and RhoC activation was impaired in TEM4-depleted cells. Finally, we showed that TEM4 and RhoC antagonize myosin II-dependent cellular contractility and the suppression of myosin II activity rescued the persistence of cellular migration of TEM4-depleted cells. Our data implicate TEM4 as an essential regulator of the actin cytoskeleton that ensures proper membrane protrusion at the leading edge of migrating cells and efficient cellular migration via suppression of actomyosin contractility.

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