ADAPTIVE MUTATIONS IN ESCHERICHIA-COLI AS A MODEL FOR THE MULTIPLE MUTATIONAL ORIGINS OF TUMORS

The cells in most tumors are found to carry multiple mutations; howeve r, based upon mutation rates determined by fluctuation tests, the freq uency of such multiple mutations should be so low that tumors are neve r detected within human populations, Fluctuation tests, which determin e the cell-division-dependent mutation rate per cell generation in gro wing cells, may not be appropriate for estimating mutation rates in no ndividing or very slowly dividing cells, Recent studies of time-depend ent, ''adaptive'' mutations in nondividing populations of microorganis ms suggest that similar measurements may be more appropriate to unders tanding the mutation origins of tumors, Here I use the ebgR and ebgA g enes of Escherichia coli to measure adaptive mutation rates where mult iple mutations are required for rapid growth, Mutations in either ebgA or ebgR allow very slow growth on lactulose (4-O-beta-D-galactosyl-D- fructose), with doubling times of 3.2 and 17.3 days, respectively. How ever, when both mutations are present, cells can grow rapidly with dou bling times of 2.7 hr, I show that during prolonged (28-day) selection for growth on lactulose, the number of lactulose-utilizing mutants th at accumulate is 40,000 times greater than can be accounted for on the basis of mutation rates measured by fluctuation tests, but is entirel y consistent with the time-dependent adaptive mutation rates measured under the same conditions of prolonged selection.