Keratins modulate hepatic cell adhesion, size and G1/S transition.
Keratins (Ks) are the intermediate filament (IF) proteins of epithelial cells. Hepatocyte IFs are made solely of keratins 8 and 18 (K8/K18), the hallmark of all simple epithelia. While K8/K18 are essential for maintaining structural integrity, there is accumulating evidence indicating that they also exert non-mechanical functions. We have reported recently that K8/K18-free hepatocytes from K8-null mice are more sensitive to Fas-mediated apoptosis, in line with an increased Fas density at the cell surface and an altered c-Flip regulation of the anti-apoptotic ERK1/2 signaling pathway. In the present study, we show that K8-null hepatocytes attach more rapidly but spread more slowly on a fibronectin substratum and undergo a more efficient G1/S transition than wild-type hepatocytes. Moreover, plectin, an IF associated protein, receptor for activated C kinase 1 (RACK1), a plectin partner, and vinculin, a key component of focal adhesions, distribute differently in spreading K8-null hepatocytes. Cell seeding leads to no differential activation of ERK1/2 in WT versus K8-null hepatocytes, whereas a stronger Akt activation is detected in K8-null hepatocytes. Insulin stimulation also leads to a differential Akt activation, implying altered Akt signaling capacity as a result of the K8/K18 loss. In addition, a delayed autophosphorylation of FAK, a target for integrin beta1 signaling, was obtained in seeding K8-null hepatocytes. These alterations in cell cycle-related events in hepatocytes in primary culture are also found in a K8-knockdown H4-II-E-C3 rat hepatoma cell line. Besides, K8/K18-free cells are smaller and exhibit a reduced rate of protein synthesis. In addition, a distinctive cyclin interplay is observed in these K8/K18-free hepatic cells, namely a more efficient cyclin A-dependent G1/S phase transition. Furthermore, K8 re-expression in these cells, following transfer of a human K8 cDNA, restores proper cell size, spreading and growth. Together, these results suggest new interrelated signaling roles of K8/18 with plectin/RACK1 in the modulation of cell attachment/spreading, size/protein synthesis and G1/S transition.
Galarneau
Luc
L
Centre de Recherche en Cancérologie, Quebec City, QC, Canada G1R 2J6.
Loranger
Anne
A
Gilbert
Stéphane
S
Marceau
Normand
N
D000818:Animals; D001483:Base Sequence; D002448:Cell Adhesion; D045744:Cell Line, Tumor; D048429:Cell Size; D002478:Cells, Cultured; D017931:DNA Primers / Q000235:genetics; D051417:Focal Adhesion Kinase 1 / Q000378:metabolism; D016193:G1 Phase; D022781:Hepatocytes / Q000166:cytology* / Q000378:metabolism*; D006801:Humans; D019012:Integrin beta1 / Q000378:metabolism; D053538:Keratin-18 / Q000378:metabolism; D053533:Keratin-8 / Q000172:deficiency / Q000235:genetics / Q000378:metabolism; D007633:Keratins / Q000378:metabolism*; D051379:Mice; D018345:Mice, Knockout; D009479:Neuropeptides / Q000378:metabolism; D051190:Plectin / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D051381:Rats; D000075370:Receptors for Activated C Kinase; D016196:S Phase; D015398:Signal Transduction; D016596:Vinculin / Q000378:metabolism
D017931:DNA Primers; D019012:Integrin beta1; D053538:Keratin-18; D053533:Keratin-8; C486595:Krt8 protein, mouse; D009479:Neuropeptides; D051190:Plectin; C090294:RACK1 protein, mouse; D000075370:Receptors for Activated C Kinase; D016596:Vinculin; D007633:Keratins; D051417:Focal Adhesion Kinase 1; C496051:Ptk2 protein, mouse; D051057:Proto-Oncogene Proteins c-akt