REEPing the harvest of reticulophagy and nucleophagy

ABSTRACT

Under stress conditions, the endoplasmic reticulum and nucleus undergo turnover through selective macroautophagy/autophagy processes termed reticulophagy and nucleophagy, respectively. Our recent study has identified the protein Hva22/Rop1/Yep1, a member of the REEP1–REEP4 subfamily of the REEP protein family, as an essential factor for both processes in the fission yeast Schizosaccharomyces pombe. In the absence of Hva22/Yep1, reticulophagy and nucleophagy cargos without surrounding autophagic membranes accumulate in the cytoplasm. Interestingly, human proteins in the REEP1–REEP4 subfamily can functionally substitute for Hva22/Yep1 to facilitate reticulophagy. Phylogenetic and synteny analyses further reveal that the budding yeast reticulophagy receptor Atg40 is also a REEP1–REEP4 subfamily member. Similar to human REEP1–REEP4 subfamily proteins, Atg40 can functionally replace Hva22/Yep1. Based on our findings, we propose that promoting reticulophagy is a conserved function of REEP1–REEP4 subfamily proteins.

The endoplasmic reticulum (ER) is a complex membrane organelle that plays a vital role in various cellular processes. In yeasts, the ER consists primarily of the nuclear envelope and cortical ER. Under stress conditions, cells activate a selective form of autophagy known as reticulophagy (or ER-phagy) to remove portions of the nuclear envelope and cortical ER. During autophagic turnover of the nuclear envelope, a portion of the nucleoplasm may undergo simultaneous autophagy, referred to as nucleophagy.

Like other selective autophagy processes, reticulophagy and nucleophagy rely on specialized cargo selectivity factors known as autophagy receptors. Autophagy receptors establish a connection between the core autophagy protein Atg8 and the cargo, thereby facilitating the sequestration of the cargo by the phagophore. In the fission yeast Schizosaccharomyces pombe, the protein Epr1 serves as an autophagy receptor for both reticulophagy and nucleophagy. In the budding yeast Saccharomyces cerevisiae, Atg39 acts as an autophagy receptor for nucleophagy, while Atg40 functions as an autophagy receptor for reticulophagy targeting the cortical ER. A large number of reticulophagy receptors have been identified in mammals.

Although reticulophagy receptors have been extensively studied, less is known about other factors required specifically for reticulophagy. In a recently published study, we conducted an imaging-based chemical mutagenesis screen in S. pombe, which led to the identification of the protein Hva22/Yep1 as an essential factor for reticulophagy and nucleophagy but not for bulk autophagy [1].

Hva22/Yep1 is a 166-amino-acid protein belonging to the REEP protein family, whose members are known to contribute to ER morphology maintenance. Like other REEP family proteins, Hva22/Yep1 localizes to the ER and possesses an ER-shaping ability. Hva22/Yep1 can self-interact through its N-terminal 113 residues, which are predicted to form three transmembrane helices and two short cytosolic helices. Interestingly, while the N-terminal 113 residues are sufficient for the ER-shaping ability, they are not sufficient for the reticulophagy function. The remaining 53 residues, predicted to form two amphipathic helices, play a crucial role in the reticulophagy function.

During the initial stage of reticulophagy and nucleophagy in fission yeast, the autophagy receptor Epr1 forms punctate structures by co-assembling with Atg8. This process occurs independently of Hva22/Yep1. In the absence of Hva22/Yep1, reticulophagy/nucleophagy cargos lacking surrounding autophagic membranes accumulate in the cytoplasm. Interestingly, in hav22/yep1Δ cells, the inner membranes and contents of the accumulated nucleophagy cargos are completely separated from their source compartment, while the outer membranes of these cargos remain connected to the ER network/nuclear envelope. This phenotype suggests that Hva22/Yep1 may play a role in the scission of the outer membranes of nucleophagy cargos (Figure 1).

Figure 1.

Schematic depicting how reticulophagy/nucleophagy fails in the absence of Hva22/Yep1. Hva22/Yep1 is indispensable for the phagophore enclosure of cargos during reticulophagy and nucleophagy. In wild-type cells, reticulophagy/nucleophagy cargos are sequestered within autophagosomes after their separation from the source membrane compartments. In the absence of Hva22/Yep1, the recruitment of the autophagic machinery at the early phase of reticulophagy/nucleophagy occurs normally, but the outer membrane of reticulophagy/nucleophagy cargos fail to separate from the source membrane compartments, resulting in the accumulation of ER- and nucleus-derived membrane structures without surrounding autophagosomes in the cytoplasm.

The REEP protein family consists of two subfamilies, the REEP1–REEP4 subfamily and the REEP5-REEP6 subfamily. In animals and fungi, it is common for each species to have proteins from both subfamilies. For instance, fission yeast species have Hva22/Yep1 from the REEP1–REEP4 subfamily and Yop1 from the REEP5-REEP6 subfamily. Interestingly, budding yeast species do not possess an obvious member of the REEP1–REEP4 subfamily. However, our phylogenetic and synteny analyses suggest that the budding yeast reticulophagy receptor Atg40 is a divergent member of the REEP1–REEP4 subfamily. Supporting the idea of a conserved role of REEP1–REEP4 subfamily proteins in promoting reticulophagy, we demonstrate that human REEP1–REEP4 subfamily proteins and budding yeast Atg40, but not human REEP5-REEP6 subfamily proteins, can support reticulophagy in hva22/yep1Δ cells. Further investigations are needed to uncover the mechanistic details of this conserved function of REEP1–REEP4 subfamily proteins in reticulophagy.

Funding Statement

We are grateful to the intramural funding from the National Institute of Biological Sciences, Beijing, and the Tsinghua Institute of Multidisciplinary Biomedical Research, Tsinghua University to L.-L.D.

Disclosure statement

No potential conflict of interest was reported by the author(s).