Short term binding of both Balb/c derived 3T3 cells and SV3T3 cells to 2B-Sepharose coated with Lens culinaris lectin (LCL) was compared with their LCL-induced agglutinability in relation to culture density and age. The cells were grown for 4 days either to low (LD) or to high density (HD); for certain experiments HD-cells were stimulated (HDS) by a short trypsin treatment and/or by addition of fresh medium. HD-3T3 cells bound somewhat faster to immobilized LCL than LD-3T3 cells, although LD-3T3 cells agglutinated at lower LCL concentrations. In the case of SV 3T3 cells, binding was much less pronounced for HD than for LD cells. The agglutinability of HD-SV 3T3 cells, however, was greater than that of the other transformed groups. Upon stimulation, binding and agglutinability data of both HD cell lines began to resemble the results obtained with LD-cells. Taken together, the date revealed in all cases an inverse relationship between binding and agglutinability, i.e., high agglutinability was closely correlated with slower binding and vice versa. The results indicate that culture density and age-dependent differences in cell surface architecture can be detected by short term binding to immobilized LCL.

Agglutination↗

PubMedDate not supplied

Protein kinases in HeLa cells and in human cervix carcinoma.

Extracts of HeLa cell fractions were analyzed by DEAE- and phospho-cellulose chromatography for their range of cyclic AMP-dependent and -independent protein kinase activities phosphorylating histone and/or phosvitin; extractions were by phosphate buffered saline (soluble protein kinases) and the non-ionic detergent NP-40 (membrane-bound protein kinases). The soluble fraction contained (i) cyclic AMP-dependent histone kinases type I and II as evidenced by their behaviour on DEAE-cellulose and inhibition by specific heat- and acid-stable protein kinase inhibitor (PKI) in a dose-related manner; both types I and II as well as their purified catalytic subunit also phosphorylated protamine and - with very low efficiency - casein but not phosvitin; (ii) a histone kinase (H), insensitive to cyclic AMP and PKI, also accepting protamine as substrate but not either casein or phosvitin; (iii) a phosvitin kinase (P), insensitive to cyclic AMP and PKI, which also phosphorylates casein but not histone or protamine. These four enzyme species were also found in NP-40 extracts of 27000 x g residues which, however, contained further histone and phosvitin kinase activities as yet unspecified. NP-40 extracts of the microsomal fraction possessed, besides unspecified histone and phosvitin kinase activity, only the phosvitin kinase P and appeared to be devoid of histone kinases I, II, and H. The occurrence and ratios of the protein kinases classified suggest an ordered distribution over the diverse subcellular fractions of HeLa cells. The overall pattern of soluble and membrane-bound histone and phosvitin kinases in extracts of cervix carcinoma tissue, the in vivo correlate of HeLa cells, closely resembled that of similar extracts of HeLa cells. HeLa cells hence appear, despite their long in vitro history, to express protein kinase activities similar to those of their in vivo ancestors, recommending them as a subject for the study of (certain) human protein kinase systems.

Chromatography, DEAE-Cellulose↗