Abstract
Human deoxycytidine deaminase APOBEC3A (Apo3A) acts as an HIV-1 restriction factor in cells of myeloid lineage yet functions separately as a potent mutator for genomic DNA. Apo3A activity and C motif deamination specificity exhibit a striking dependence on pH that reflects these two distinct biological processes. Upon infection of macrophages, HIV-1 induces the formation of autophagosomes, and requires autophagosomes for replication, whereas inhibiting lysosomal fusion indicative of late stage autophagy. Here we show that Apo3A has optimal activity and a strict 5'-YYCR motif specificity in the pH 5.8-6.1 range, characteristic of enclosed autophagosomal membrane compartments, and reflective of the mutation pattern of HIV-1. In contrast to the high activity and narrow specificity of Apo3A at acid pH, a 13-30-fold reduction in specific activity is accompanied by relaxed C deamination specificity at pH 7.4-8. Notably, Apo3A is also expressed in keratinocytes, and is up-regulated in skin lesions. At pH 7.9, we show that Apo3A generates transcription-dependent CC → TT tandem mutations on the non-transcribed strand, a hallmark signature of skin cancer. The biochemical data taken in conjunction with the biological up-regulation of Apo3A in skin lesions suggests that enzyme-catalyzed deaminations at adjacent C sites followed by normal replication generating CC → TT mutations provides an alternative molecular basis for the initiation events in skin cancer in contrast to well established pathways in which CC dimers formed in response to UV radiation either undergo nonenzymatic spontaneous deaminations or aberrant replication.
Highlights
Apo3A restricts HIV-1 in myeloid cells and mutates genomic DNA
The pH-dependent catalytic properties of Apo3A reflect targeting of HIV-1 cDNA and “off targeting” of genomic DNA
The biochemical data taken in conjunction with the biological up-regulation of Apo3A in skin lesions suggests that enzyme-catalyzed deaminations at adjacent C sites followed by normal replication generating CC 3 TT mutations provides an alternative molecular basis for the initiation events in skin cancer in contrast to well established pathways in which CC dimers formed in response to UV radiation either undergo nonenzymatic spontaneous deaminations or aberrant replication
Summary
Apo3A restricts HIV-1 in myeloid cells and mutates genomic DNA. Results: Optimal Apo3A activity and narrow deamination specificity occur at acidic pH; weak activity and broad specificity, featuring CC 3 TT mutations, occur at physiological pH. Human deoxycytidine deaminase APOBEC3A (Apo3A) acts as an HIV-1 restriction factor in cells of myeloid lineage yet functions separately as a potent mutator for genomic DNA. APOBEC proteins require tight cellular regulation to prevent C deaminations in DNA from occurring at the wrong place or time; “off targeted” mutations by AID can cause B-cell lymphoma [32, 33], whereas inadvertent action of Apo3B has recently been implicated in breast cancer [30]. A biochemical analysis of Apo3A activity and motif deamination specificity at acidic pH, corresponding to enclosed autophagosomal membrane compartments, and at physiological pH, corresponding to cell nuclei, indicates the presence of a dual mutagenic mechanism that might link disparate HIV-1 inactivation in myeloid cells and susceptibility to skin cancer in precancerous skin lesions
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