Influence of proliferative rates and A system substrate availability on proline transport in primary cell cultures of the R3230AC mammary tumor

J Cell Physiol. 1980 Nov;105(2):287-300. doi: 10.1002/jcp.1041050212.

Abstract

Regulation of A system amino acid transport was studied in primary cultures of the R3230AC mammary adenocarcinoma. Higher rates of carrier-mediated Na+-dependent proline transport, vc, was decreased and was attributed to a two-fold decrease in Vmax and a two-fold increase in Km. When compared to cells grown in standard media (Eagle's minimal essential medium, MEM), cells grown in media supplemented with A system substrates (alanine, serine, glycine, and proline) demonstrated adaptive decreases in proline transport; the decrease was due to two-fold reduction in Vmax, with no change in Km for proline. Even in the presence of preferred substrates for the A system, a density-dependent decrease in proline transport was manifested. Both fast- and slow-growing cultures maintained in MEM exhibited rapid increases in proline transport when switched to buffers devoid of amino acids; two-fold increases in Vmax were seen within 4 hr, but Km was unchanged. This starvation-induced adaptation was completely prevented by inclusion in the buffer of 10 mM proline, 0.1 mM alpha-(methylamino)-isobutyric acid (MetAIB) or 10 mM serine, whereas inclusion of the poorer A system substrate, phenylalanine (10 mM), had no effect. The effects of MetAIB to prevent starvation-induced increases in proline transport were dose-related, rapid, and reversible. Amino acid starvation-induced increases in proline transport were partially blocked by cycloheximide or actinomycin D. Data were obtained demonstrating a temporal relationship between increasing intracellular [proline] and decreasing vc for proline uptake. In addition, efflux of proline from preloaded cells preceded the increase in initial rates of proline entry. Taken together, we concluded that: 1) A system transport in primary cultures of this mammary adenocarcinoma is regulated by cell density as well as by availability of A system substrates, but these two types of regulation are kinetically distinct; and 2) starvation-induced enhancement of proline transport appears to be due to release from transinhibition, but may also involve a derepression-repression type of mechanism.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acids / metabolism
  • Animals
  • Biological Transport, Active
  • Cell Division*
  • Cells, Cultured
  • Kinetics
  • Mammary Neoplasms, Experimental
  • Neoplasm Proteins / biosynthesis
  • Proline / metabolism*
  • RNA, Neoplasm / biosynthesis
  • Rats
  • Sodium / pharmacology

Substances

  • Amino Acids
  • Neoplasm Proteins
  • RNA, Neoplasm
  • Proline
  • Sodium