PROTEIN AND LIPID LATERAL DIFFUSION IN NORMAL AND ROUS-SARCOMA VIRUS-TRANSFORMED CHICK-EMBRYO FIBROBLASTS

We measured the lateral diffusion of the fluorescent lipid analogue dioctadecylindocarbocyanine iodide (Dil) and of membrane glycoproteins labeled with tetramethylrhodamine (TRITC) succinyl concanavalin A (SConA) via fluorescence photobleaching recovery (FPR) at selected times during a temperature downshift experiment on transformation-defective temperature-sensitive (td-ts) Rous sarcoma virus (RSV) NY68-transformed chicken embryo fibroblasts (CEF) and on identically treated CEF and RSV-transformed CEF. There were no significant differences in the lateral diffusion of Dil at any of the times measured. The lateral diffusion of TRITC-SConA on the RSV-transformed CEF, (1.32 +/- 0.12). 10(-10) cm2 s-1, was approximately two times faster than that observed in normal CEF, (0.61 +/- 0.06) . 10(-10) cm2 s-1. In the cells undergoing RSV NY68-mediated transformation, TRITC-SConA diffusion increased over a 24-h period from a value comparable to that observed in normal CEF, (0.72 +/- 0.13) . 10(-10) cm2 s-1 to a value comparable to thc RSV-CEF transformed cells, (1.74 +/- 0.20) . 10(-10) cm2 s-1. All diffusion measurements reported were made at the permissive temperature for RSV-NY68 (35-degrees-C) unless stated otherwise. The changes in the lateral diffusion of TRITC-SConA occurred between the fifth and twelfth hour of the downshift course and could be associated with cytoskeletal disruption and/or fibronectin degradation, both known to occur at this time in RSV-transformed cells. To assess the contribution of extracellular matrix (ECM) degradation, SConA mobility was measured in normal and RSV-transformed cells treated with trypsin. This treatment increased SConA mobility approximately 4-fold in the normal cells relative to untreated controls and only 2-fold in the RSV-CEF transformed cells. No significant difference in SConA mobility between trypsinized spherical normal and transformed cells was apparent.