Molecular and structural mechanisms of ZZ domain-mediated cargo recognition by autophagy receptor Nbr1

Abstract

In selective autophagy, cargo selectivity is determined by autophagy receptors. However, it remains scarcely understood how autophagy receptors recognize specific protein cargos. In the fission yeast Schizosaccharomyces pombe, a selective autophagy pathway termed Nbr1-mediated vacuolar targeting (NVT) employs Nbr1, an autophagy receptor conserved across eukaryotes including humans, to target cytosolic hydrolases into the vacuole. Here, we identify two new NVT cargos, the mannosidase Ams1 and the aminopeptidase Ape4, that bind competitively to the first ZZ domain of Nbr1 (Nbr1-ZZ1). High-resolution cryo-EM analyses reveal how a single ZZ domain recognizes two distinct protein cargos. Nbr1-ZZ1 not only recognizes the N-termini of cargos via a conserved acidic pocket, similar to other characterized ZZ domains, but also engages additional parts of cargos in a cargo-specific manner. Our findings unveil a single-domain bispecific mechanism of autophagy cargo recognition, elucidate its underlying structural basis, and expand the understanding of ZZ domain-mediated protein-protein interactions.

ZZ domains keep cytosol to vacuole delivery whiZZing along

“…In order to further characterize the mechanism of binding, Wang et al (2021) determined the cryo‐EM structure of Nbr1's ZZ1 domain in complex with Ams1 or Ape4. The resulting structures revealed two binding interfaces that work cooperatively, one which is common to ZZ domain‐containing proteins such as p62/SQSTM1, p300, ZZZ3, and HERC2 (Zhang et al , 2019), and the other one being cargo‐specific.…”

“…In this issue, Wang et al (2021) performed a mass spectrometry‐based protein interactome analysis using Nbr1 as bait and identified Ams1 and Ape4, homologs of the Cvt pathway cargos in the budding yeast, as novel cargos for the NVT pathway in the fission yeast. The fission yeast Nbr1 has three ZZ domains (ZZ1–3).…”

“…The UBA domain, which is lacking in S. pombe Nbr1, interacts with polyubiquitin and therefore mediates the degradation of ubiquitinated proteins (Johansen & Lamark, 2020). On the other hand, Wang et al (2021) found that the ZZ domain is involved in the biosynthesis of hydrolases in the fission yeast (Liu et al , 2015; Wang et al , 2021). Thus, it might be hypothesized that the ZZ and UBA domains mediate biosynthetic and degradation pathways, respectively.…”

“…The adaptor proteins p62/SQSTM1, Nbr1, and Atg19/Atg34 belong to the same family. The Cvt pathway transports Ams1 and Ape4 via macroautophagy-like mechanisms, whereas the NVT pathway delivers Lap2-Ape2, Ams1, and Ape4 (the latter two are newly identified cargos in Wang et al (2021)) through microautophagy-like mechanisms. (B) Domain structures of human p62/SQSTM1, human NBR1, Schizosaccharomyces pombe Nbr1, Saccharomyces cerevisiae Atg19 and Atg34, and cargo proteins recognized by each domain.…”

“…Selective autophagy relies on adaptor proteins to bind and transport cargos (or substrates) to the lysosome or vacuole, yet the mechanisms for cargo recognition are not well understood. In this issue, Wang et al (2021) showed that in the fission yeast, Nbr1, a homolog of a mammalian selective autophagy adaptor, recognizes vacuolar hydrolases Ams1 and Ape4 through both versatile and cargo-specific interactions with the Nbr1 ZZ1 domain. A utophagy is a general term describing processes that deliver cytoplasmic components to the lysosome or vacuole for degradation.…”