Regulation of H2a-specific proteolysis by the histone H3:H4 tetramer.
We have studied the limited cleavage of H2a in the H2a:H2b histone dimer by the H2a-specific protease under physiological conditions (neutral pH, 0.1 M NaCl) using a variety of histone-DNA reconstitutes as substrates and/or regulators of the partially purified enzyme. Under these conditions the protease cleaves H2a in "native" dimer-DNA reconstitutes but not in "native" octamer-DNA reconstitutes. Treatment of the enzyme with saturating amounts of H3:H4 tetramer-DNA prior to addition of dimer-DNA substrate results in complete inhibition of H2a-specific proteolysis. Sucrose gradient sedimentation experiments indicate that the protease binds reversibly to tetramer-DNA and that this leads to the reversible inhibition of enzymatic activity. Using three different tetramer-DNA complexes, we found native tetramer-DNA to be a more effective inhibitor than either trypsin-treated tetramer-DNA or acetylated tetramer-DNA. We conclude that under physiological conditions, the H2a-specific protease binds primarily to the highly basic amino-terminal domain of the H3:H4 tetramer, and this binding lowers the effective concentration of enzyme available to cleave H2a. Although no cleaved H2a is produced when protease is mixed with native octamer-DNA, incubation of the enzyme with acetylated octamer-DNA results in H2a-specific proteolysis. This is the first demonstration that the H2a-specific protease activity can be modulated by a physiologically relevant process (e.g. histone acetylation). We propose that the sequestered protease may be functionally regulated in vivo through reversible post-translational modifications to the NH2-terminal domains of the histone H3:H4 tetramer.
Elia
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MC
Department of Biology, Johns Hopkins University, Baltimore, Maryland 21218.
Moudrianakis
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EN
D000107:Acetylation; D000818:Animals; D002417:Cattle; D002499:Centrifugation, Density Gradient; D004247:DNA / Q000378:metabolism* / Q000494:pharmacology; D010450:Endopeptidases / Q000378:metabolism*; D006657:Histones / Q000378:metabolism* / Q000494:pharmacology; D007700:Kinetics; D046911:Macromolecular Substances; D009994:Osmolar Concentration; D011480:Protease Inhibitors; D011499:Protein Processing, Post-Translational; D014357:Trypsin / Q000494:pharmacology
D006657:Histones; D046911:Macromolecular Substances; D011480:Protease Inhibitors; D004247:DNA; D010450:Endopeptidases; D014357:Trypsin; C021361:histone proteases