ZCCHC3 is a stress granule zinc knuckle protein that strongly suppresses LINE-1 retrotransposition

Retrotransposons have generated about half of the human genome and LINE-1s (L1s) are the only autonomously active retrotransposons. The cell has evolved an arsenal of defense mechanisms to protect against retrotransposition with factors we are only beginning to understand. In this study, we investigate Zinc Finger CCHC-Type Containing 3 (ZCCHC3), a gag-like zinc knuckle protein recently reported to function in the innate immune response to infecting viruses. We show that ZCCHC3 also severely restricts human retrotransposons and associates with the L1 ORF1p ribonucleoprotein particle. We identify ZCCHC3 as a bona fide stress granule protein, and its association with LINE-1 is further supported by colocalization with L1 ORF1 protein in stress granules, dense cytoplasmic aggregations of proteins and RNAs that contain stalled translation pre-initiation complexes and form when the cell is under stress. Our work also draws links between ZCCHC3 and the anti-viral and retrotransposon restriction factors Mov10 RISC Complex RNA Helicase (MOV10) and Zinc Finger CCCH-Type, Antiviral 1 (ZC3HAV1, also called ZAP). Furthermore, collective evidence from subcellular localization, co-immunoprecipitation, and velocity gradient centrifugation connects ZCCHC3 with the RNA exosome, a multi-subunit ribonuclease complex capable of degrading various species of RNA molecules and that has previously been linked with retrotransposon control. Author summaryRetrotransposons are mobile DNA elements that duplicate themselves by a "copy and paste" mechanism using an RNA intermediate. Their misregulation has been linked with some cancers, neuropathologies, and cellular aging. Consequently, the cell has evolved a battery of defenses to protect against retrotransposition. The ancient origin of endogenous retrotransposons suggests their inhibition may have been an evolutionary driver for some host restriction factors, and these may later have been co-opted for defense against exogenous viruses. Zinc Finger CCHC-Type Containing 3, ZCCHC3, protein is a putative host cell restriction factor whose antiviral and innate immune response actions have recently been identified, and that we here show interacts with and limits activity of human retrotransposons in cell culture. In this work we also draw connections between ZCCHC3 and the RNA helicase MOV10 and zinc-finger protein ZAP, host proteins known to restrict viral infection in mammals and previously shown by ourselves and others to potently inhibit human retrotransposition. We also consider that ZCCHC3 may be functionally associated with the cytoplasmic RNA exosome as well as cytoplasmic stress granules, membraneless ribonucleoprotein aggregates that are important for both mRNA metabolism and viral infectivity. Our experiments improve understanding of ZCCHC3 restriction function and may in the future have potential application to the therapeutic treatment of human infectious diseases.