ZNK1 senses zinc and degrades zinc transporter mRNA in trypanosomes

Abstract Like other cells, parasitic and other trypanosomatids sense and regulate Zn 2+ transport, but the mechanisms involved remained unknown. Here we identify a trypanosome RNA-binding protein which specifically eliminates ZIP3 Zn 2+ -transporter mRNA in Zn 2+ -replete conditions. We first demonstrated that Trypanosoma brucei ZIP3 mRNA abundance is subject to 3’-untranslated region (3’-UTR) and Zn 2+ -dependent negative control. A genome-wide RNA interference library screen, using a reporter associated with the ZIP3 3’-UTR, identified Tb927.11.9510 as a candidate Zn 2+ -sensor. We name this protein Zinc Nuclear Knuckles 1 (ZNK1) since it localises to the nucleus and contains several Zn 2+ -knuckle motifs. ZNK1 is conserved among trypanosomatids, and a PIN-domain suggests a ribonuclease-based mechanism. We validate ZNK1 as a ZIP3 3’-UTR dependent negative regulator and identify a GU-repeat motif in the ZIP3 3’-UTR that is predictive of negative control by ZNK1. We use Cas9-editing to knockout ZNK1, and RNA-seq to assess the consequences, revealing highly specific accumulation of ZIP3 transcripts in znk1 -null cells. We conclude that ZNK1 senses Zn 2+ -abundance and eliminates ZIP3 mRNA in a Zn 2+ -dependent manner. We suggest that trypanosomatid ZNK1 is an RNA-specific zinc finger nuclease that binds ZIP3 3’-UTRs and degrades ZIP3 mRNA only when the tandem sensor modules are coordinated with Zn 2+ . Key points Trypanosome Zinc Nuclear Knuckles 1 (ZNK1) is a zinc-sensor that eliminates zinc transporter mRNA. ZNK1 negative control operates via the transporter mRNA 3’-untranslated region. The findings indicate that trypanosomatid ZNK1 is a conserved RNA-specific zinc finger nuclease.