In 2012, two separate studies reported that FLCN interacts with Plakophilin4 (PKP4) to regulate RhoA signalling and cell-cell adhesion. PKP4, together with Ect2, interacts with RhoA to regulate Rho signalling, contractile ring formation and contraction, and actin reorganisation during cell division (Keil et al., 2007; Keil et al., 2009; Wolf et al., 2006). PKP4 is also thought to regulate cell-cell adhesion as it localises to cell-cell junctions, particularly desmosomes and adherens junctions (Hatzfield et al., 2003), and is one of a family of p120 catenins that regulate the turnover of cadherin proteins at the plasma membrane (Hatzfield, 2005). This month, researchers published results characterising how PKP4 might control cell-cell adhesion.
Using a yeast two-hybrid screen, Keil and Hatzfield (2014) show that PKP4 interacts with the small GTPase Rab11a, with particular affinity for active Rab11a. Bi-molecular fluorscence complementation experiments in HeLa and MCF-7 cells show punctate co-localisation of PKP4 and Rab11a throughout the cytoplasm, with a few cells showing peri-nuclear localisation of the two proteins, suggesting that this interaction occurs in vivo.
siRNA knockdown of PKP4 led to an increase in the proportion of cells showing perinuclear accumulation of Rab11a. Inhibition of RhoA signalling using the ROCK inhibitor Y27632 showed a similar pattern of Rab11a mislocalisation, suggesting that PKP4 likely controls Rab11a localisation via the Rho signalling pathway. Interestingly, when cells were treated with nocodazole, thus ablating microtubule formation, Rab11a became predominantly localised at cell-cell contacts. This suggests that Rab11a is present at cell-cell junctions, and the protein is transported from the plasma membrane to the nucleus via microtubule associated transport proteins, and is recycled to the plasma membrane by PKP4.
Rab11a is a small GTPase that, together with Rab4 and Rab5, regulates endocytosis and the recycling of proteins to the plasma membrane, including the cell junction protein E-cadherin. siRNA knockdown of Rab11a in HeLa cells increased the amount of PKP4 localised at cell-cell junctions, suggesting that Rab11a regulates the localisation of PKP4 at the plasma membrane. Increased levels of PKP4 at cell-cell junctions increased cell-cell contact strength, as shown by shearing experiments. This phenotype was reversed by knocking down PKP4, suggesting that PKP4 localisation and turn over at cell junctions directly affects cell-cell adhesion.
Together these data suggest that Rab11a and PKP4 regulate each others’ localisation and turn over at the plasma membrane, which ultimately regulates how strongly the cell adheres to its neighbours. As FLCN is a confirmed interacting partner of PKP4 (Medvetz et al., 2012; Nahorski et al., 2012), it would be interesting to determine whether FLCN also converges on the same pathway as PKP4 and Rho signalling to regulate Rab11a function, and whether Rab11a controls FLCN localisation in turn. Indeed, similarly to Rab11a, FLCN shows a punctate pattern of co -localisation with PKP4 throughout the cytoplasm, and localises to cell junctions, making it seem plausible that the two might interact.
Furthermore, a study published in 2010 systematically tested the GAP activity of known DENN domain proteins toward multiple Rab proteins (Yoshimura et al., 2010), and no known activating protein was found for Rab11a. In 2012, FLCN was the first reported protein carrying a non-canonical DENN domain, and to have activity towards Rab35 in vitro (Nookala et al., 2012). Thus it is tempting to speculate that FLCN may activate Rab11a via its interaction with PKP4, and control Rho signalling and cell-cell adhesion.
- Hatzfeld M, Green KJ, & Sauter H (2003). Targeting of p0071 to desmosomes and adherens junctions is mediated by different protein domains. Journal of cell science, 116 (Pt 7), 1219-33 PMID: 12615965
- Hatzfeld M (2005). The p120 family of cell adhesion molecules. European journal of cell biology, 84 (2-3), 205-14 PMID: 15819401
- Keil R, Wolf A, Hüttelmaier S, & Hatzfeld M (2007). Beyond regulation of cell adhesion: local control of RhoA at the cleavage furrow by the p0071 catenin. Cell cycle (Georgetown, Tex.), 6 (2), 122-7 PMID: 17264675
- Keil R, Kiessling C, & Hatzfeld M (2009). Targeting of p0071 to the midbody depends on KIF3. Journal of cell science, 122 (Pt 8), 1174-83 PMID: 19339549
- Keil R, & Hatzfeld M (2014). The armadillo protein p0071 is involved in Rab11-dependent recycling. Journal of cell science, 127 (Pt 1), 60-71 PMID: 24163434
- Medvetz DA, Khabibullin D, Hariharan V, Ongusaha PP, Goncharova EA, Schlechter T, Darling TN, Hofmann I, Krymskaya VP, Liao JK, Huang H, & Henske EP (2012). Folliculin, the product of the Birt-Hogg-Dube tumor suppressor gene, interacts with the adherens junction protein p0071 to regulate cell-cell adhesion. PloS one, 7 (11) PMID: 23139756
- Nahorski MS, Seabra L, Straatman-Iwanowska A, Wingenfeld A, Reiman A, Lu X, Klomp JA, Teh BT, Hatzfeld M, Gissen P, & Maher ER (2012). Folliculin interacts with p0071 (plakophilin-4) and deficiency is associated with disordered RhoA signalling, epithelial polarization and cytokinesis. Human molecular genetics, 21 (24), 5268-79 PMID: 22965878
- Nookala RK, Langemeyer L, Pacitto A, Ochoa-Montaño B, Donaldson JC, Blaszczyk BK, Chirgadze DY, Barr FA, Bazan JF, & Blundell TL (2012). Crystal structure of folliculin reveals a hidDENN function in genetically inherited renal cancer. Open biology, 2 (8) PMID: 22977732
- Wolf A, Keil R, Götzl O, Mun A, Schwarze K, Lederer M, Hüttelmaier S, & Hatzfeld M (2006). The armadillo protein p0071 regulates Rho signalling during cytokinesis. Nature cell biology, 8 (12), 1432-40 PMID: 17115030
- Yoshimura S, Gerondopoulos A, Linford A, Rigden DJ, & Barr FA (2010). Family-wide characterization of the DENN domain Rab GDP-GTP exchange factors. The Journal of cell biology, 191 (2), 367-81 PMID: 20937701
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