Modulation of Cell Proliferation by Cytokeratins K10 and K16

Abstract

The members of the large keratin family of cytoskeletal proteins are expressed in a carefully regulated tissueand differentiation-specific manner. Although these proteins are thought to be involved in imparting mechanical integrity to epithelial cells, the functional significance of their complex differential expression is still unclear. Here we provide new data suggesting that the expression of particular keratins may influence cell proliferation. Specifically, we demonstrate that the ectopic expression of K10 inhibits the proliferation of human keratinocytes in culture, while K16 expression appears to promote the proliferation of these cells. Other keratins, such as K13 or K14, do not significantly alter this parameter. K10-induced inhibition is reversed by the coexpression of K16 but not that of K14. These results are coherent with the observed expression pattern of these proteins in the epidermis: basal, proliferative keratinocytes express K14; when they terminally differentiate, keratinocytes switch off K14 and start K10 expression, whereas in response to hyperproliferative stimuli, K16 replaces K10. The characteristics of this process indicate that K10 and K16 act on the retinoblastoma (Rb) pathway, as (i) K10-induced inhibition is hampered by cotransfection with viral oncoproteins which interfere with pRb but not with p53; (ii) K10-mediated cell growth arrest is rescued by the coexpression of specific cyclins, cyclindependent kinases (CDKs), or cyclin-CDK complexes; (iii) K10-induced inhibition does not take place in Rbdeficient cells but is restored in these cells by cotransfection with pRb or p107 but not p130; (iv) K16 efficiently rescues the cell growth arrest induced by pRb in HaCaT cells but not that induced by p107 or p130; and (v) pRb phosphorylation and cyclin D1 expression are reduced in K10-transfected cells and increased in K16-transfected cells. Finally, using K10 deletion mutants, we map this inhibitory function to the nonhelical terminal domains of K10, hypervariable regions in which keratin-specific functions are thought to reside, and demonstrate that the presence of one of these domains is sufficient to promote cell growth arrest.Keratins are a large family of proteins which form the intermediate filament (IF) cytoskeleton of epithelial cells and their appendages, hairs and nails (reviewed in references 8 and 15). These proteins are subdivided according to biochemical criteria into two subfamilies: type I, or acidic keratins, and type II, or neutral-basic keratins. This division also has important structural and functional implications, since to build up a wellorganized IF cytoskeleton, tetramers containing equimolar amounts of each keratin subtype are required. Like all IF proteins, keratins consist of a central ␣-helical rod domain responsible for dimerization and higher-order polymerization. The rod domain is flanked by globular head (amino) and tail (carboxyl) domains, the functions of which are still unclear. Variations in these nonhelical end domains largely account for the differe...

PTEN tumour suppressor is linked to the cell cycle control through the retinoblastoma protein

“…We observed that transfected pRb is e ciently phosphorylated in C33A, in parallel with an induction of cyclin D1 protein, in agreement with our previous results (Paramio et al, 1999), whilst the co-expression of PTEN inhibits this hyperphosphorylation and reduces the cyclin D1 protein expression (Figure 3d). Given that PTEN may dephosphorylate phosphatidylinositol 3,4,5 triphosphate (Maehama and Dixon, 1998), thus inhibiting the PI-3 kinase signalling cascade (Li and Sun, 1998;Stambolic et al, 1998;Furnari et al, 1998;Haas-Kogan et al, 1998;Wu et al, 1998), and the fact that PI-3K can modulate pRb phosphorylation (Prennan et al, 1997;Klippel et al, 1998), we also analysed if the co-expression of a catalitically active subunit of PI-3K (p110CAAX, kindly provided by Dr J Downward, ICRF, London) could restore the hyperphosphorylation of pRb.…”

“…To this, C33A cells were co-transfected with either pRb, p107 or p130 along with PTEN or empty vector and the number of colonies was scored in each case. It is worth mentioning that, in C33A cells pRb expression is unable to cause cell cycle arrest (93+10.5% colonies with respect to empty vector, see also Paramio et al (1999); Zhu et al (1993Zhu et al ( , 1995). We observed that the (Paramio et al, 1998;.…”

Unique and overlapping functions of pRb and p107 in the control of proliferation and differentiation in epidermis

“…4) might be of a particular relevance. Our recent work has demonstrated that this keratin may impose a cell cycle arrest in keratinocytes in a pRb-dependent manner (Paramio et al, 1999;Paramio et al, 2001a;Santos et al, 2002). Here, we show that K10-expressing cells do not exhibit a proliferative arrest.…”

Functional roles of Akt signaling in mouse skin tumorigenesis

“…These data indicate that PI-3K activity is involved in the early activation of Akt, although other mechanisms may also be implicated. We have previously demonstrated that the expression of keratin K10 inhibits keratinocyte proliferation (Paramio et al, 1999b), and more recently have associated such inhibition to the repression of Akt activities (Paramio et al, 2001). Therefore, the increased activity of Akt could also be due to the loss of K10 expression in tumors.…”