Requirement for alanine in the amino acid control of deprivation-induced protein degradation in liver.

Abstract

Protein degradation in liver is actively controlled by a small group of inhibitory amino acids-leucine, tyrosine (or phenylalanine), glutamine, proline, histidine, tryptophan, and methionine. Other evidence, however, suggests that one or more of the remaining 12 noninhibitory amino acids is also required for suppression of proteolysis at normal concentrations. This question was investigated in livers of fed rats perfused in the single-pass mode. The deletion of alanine at normal (lx), but not at 4x or 1Ox normal, plasma amino acid concentrations evoked a near-maximal acceleration of protein degradation. No other noninhibitory amino acid was effective. Because alanine alone was not directly inhibitory and its omission was not associated with a decrease in inhibitory amino acid pools, alanine was presumed to act as a coregulator in the expression of inhibitory activity. When tested alone, the inhibitory group was as effective as the complete mixture at 0.5x and 4x levels, but it lost its suppressive ability within a narrow zone of concentration centered slightly above 1x. The addition of 1x (0.48 mM) alanine completely restored the inhibition. Pyruvate and lactate could be effectively substituted, but only at concentrations 10-20 times greater than that of alanine. These, together with earlier findings, indicate the existence of a regulatory complex that recognizes specific amino acids and transmits positive and negative signals to proteolytic sites. The results also suggest that alanine can provide an important regulatory link between energy demands and protein degradation.

Utilizing flow cytometry to monitor autophagy in living mammalian cells

“…The effect of specific amino acids has been extensively studied in the past and a group of specific single amino acids was shown to inhibit autophagy-mediated proteolysis. [20][21][22][23][24] To test whether induction of autophagy by specific amino acids is detectable by FACS analysis, cells stably expressing GFP-LC3 were incubated for 5 hours in medium, systematically omitted of a specific single amino acid and the level of autophagic activity was examined using flow cytometry. As shown in Figure 4A, only specific amino acids inhibit autophagy.…”

Skeletal Muscle Proteases and Protein Turnover

“…John, 1976;Poso and Mortimore, 1984), a number of studies have now shown that other nonlysosomal proteases are also involved in metabolic turnover of intracellular proteins (Zeman et al, 1985;Hough et al, 1986;Fagan et al, 1987). The lack of definitive information on the proteases involved and the inability thus far to analyze intracellular protein degradation by monitoring intermediates in the degradation process have hampered studies on the mechanism of intracellular protein degradation.…”

Regulation of RNA degradation in cultured rat hepatocytes: Effects of specific amino acids and insulin

“…The observations on RNA degradation presented in Figure 1 showed a gradual and continuous inhibitory effectiveness of the eight amino acids known as regulators of proteolysis. In perfused livers, the inhibitory effectiveness of this regulatory group on protein degradation nearly disappeared within a narrow zone between 0.5 and four times normal plasma concentration of amino acids (Poso and Mortimore, 1984;. Such a loss of inhibition did not appear for the degradation of RNA in cultured rat hepatocytes with physiological concentration of regulatory amino acids (AA8 group).…”