The endonuclease Ankle1 requires its LEM and GIY-YIG motifs for DNA cleavage in vivo

Abstract

SummaryThe LEM domain (for lamina-associated polypeptide, emerin, MAN1 domain) defines a group of nuclear proteins that bind chromatin through interaction of the LEM motif with the conserved DNA crosslinking protein, barrier-to-autointegration factor (BAF). Here, we describe a LEM protein annotated in databases as 'Ankyrin repeat and LEM domain-containing protein 1' (Ankle1). We show that Ankle1 is conserved in metazoans and contains a unique C-terminal GIY-YIG motif that confers endonuclease activity in vitro and in vivo. In mammals, Ankle1 is predominantly expressed in hematopoietic tissues. Although most characterized LEM proteins are components of the inner nuclear membrane, ectopic Ankle1 shuttles between cytoplasm and nucleus. Ankle1 enriched in the nucleoplasm induces DNA cleavage and DNA damage response. This activity requires both the catalytic C-terminal GIY-YIG domain and the LEM motif, which binds chromatin via BAF. Hence, Ankle1 is an unusual LEM protein with a GIY-YIG-type endonuclease activity in higher eukaryotes.

Cell- and Virus-Mediated Regulation of the Barrier-to-Autointegration Factor's Phosphorylation State Controls Its DNA Binding, Dimerization, Subcellular Localization, and Antipoxviral Activity

“…These data shed further light on mechanisms employed by cellular kinase such as VRK1 or the vaccinia virus kinase B1 to regulate BAF's functions in mitosis and antiviral host defense. Finally, as phosphorylation clearly regulates BAF at multiple levels, it is tempting to speculate that it also impacts other cellular processes recently associated with BAF, such as transcriptional regulation (5,8,52,53) and the DNA damage response (6,9,54).…”

“…BAF can interact with double-stranded DNA in a sequence-independent manner, homodimerize to crossbridge DNA, and form higher-order nucleoprotein complexes (1)(2)(3)(4). BAF also interacts with many cellular proteins, including LAP2/emerin/MAN1 (LEM) domain proteins that reside in the nuclear envelope, histones, lamins, transcription factors, and DNA damage response (DDR) proteins (5)(6)(7)(8)(9)(10). Using these interactions, BAF is thought to act as a tethering protein to bring together chromatin DNA and LEM proteins during late stages of mitosis when the nuclear envelope (NE) is being reassembled.…”

A new class of ultrafine anaphase bridges generated by homologous recombination

“…The mammalian ortholog of LEM-3 is known as ANKLE1 [68,69]. LEM-3/ANKLE1 contains N-terminal Ankyrin repeats, a LEM domain and a C-terminal GIY-YIG nuclease motif that is similar to that found in SLX1 nuclease.…”

“…LEM-3/ANKLE1 contains N-terminal Ankyrin repeats, a LEM domain and a C-terminal GIY-YIG nuclease motif that is similar to that found in SLX1 nuclease. Although there is presently little known about the specificity of the LEM-3/ANKLE1 nuclease activity, it has been shown to cut both single-stranded and duplex DNA [68,69]. Based on these studies, it is conceivable that UFBs arising from unresolved replication/recombination intermediates might also be acted upon by ANKLE1.…”

The Barrier to Autointegration Factor: Interlocking Antiviral Defense with Genome Maintenance

“…Interestingly, Ankle2 can also inhibit the catalytic activity of VRK1, further enhancing BAF dephosphorylation. Ankle1, another more recently discovered BAF partner protein, possesses both a LEM domain and a GIY-YIG nuclease domain (20); it is tempting to speculate that if Ankle1 is recruited to a viral genome by BAF, Ankle1 may facilitate degradation of that DNA as a unique antiviral defense. Together, these studies suggest that BAF's role in host-virus interaction probably involves protein partners and posttranslational modification that also control BAF during mitosis and other cellular processes.…”