E Tian

C. elegans Screen Identifies Autophagy Genes Specific to Multicellular Organisms

The molecular understanding of autophagy has originated almost exclusively from yeast genetic studies. Little is known about essential autophagy components specific to higher eukaryotes. Here we perform genetic screens in C. elegans and identify four metazoan-specific autophagy genes, named epg-2, -3, -4, and -5. Genetic analysis reveals that epg-2, -3, -4, and -5 define discrete genetic steps of the autophagy pathway. epg-2 encodes a coiled-coil protein that functions in specific autophagic cargo recognition. Mammalian homologs of EPG-3/VMP1, EPG-4/EI24, and EPG-5/mEPG5 are essential for starvation-induced autophagy. VMP1 regulates autophagosome formation by controlling the duration of omegasomes. EI24 and mEPG5 are required for formation of degradative autolysosomes. This study establishes C. elegans as a multicellular genetic model to delineate the autophagy pathway and provides mechanistic insights into the metazoan-specific autophagic process.

SEPA-1 Mediates the Specific Recognition and Degradation of P Granule Components by Autophagy in C. elegans

How autophagy, an evolutionarily conserved intracellular catabolic system for bulk degradation, selectively degrades protein aggregates is poorly understood. Here, we show that several maternally derived germ P granule components are selectively eliminated by autophagy in somatic cells during C. elegans embryogenesis. The activity of sepa-1 is required for the degradation of these P granule components and for their accumulation into aggregates, termed PGL granules, in autophagy mutants. SEPA-1 forms protein aggregates and is also a preferential target of autophagy. SEPA-1 directly binds to the P granule component PGL-3 and also to the autophagy protein LGG-1/Atg8. SEPA-1 aggregates consistently colocalize with PGL granules and with LGG-1 puncta. Thus, SEPA-1 functions as a bridging molecule in mediating the specific recognition and degradation of P granule components by autophagy. Our study reveals a mechanism for preferential degradation of protein aggregates by autophagy and emphasizes the physiological significance of selective autophagy during animal development.

epg-1 functions in autophagy-regulated processes and may encode a highly divergent Atg13 homolog in C. elegans

Autophagy is an evolutionarily conserved intracellular catabolic system for degradation of long-lived proteins or damaged organelles. In this study, we have identified and characterized a new gene, epg-1, that plays a role in the autophagy pathway in C. elegans. Loss of function of epg-1 causes defects in various autophagy-regulated processes, including degradation of aggregate-prone proteins and optimal survival of animals during starvation. epg-1 encodes a novel protein that shows limited sequence similarity to the yeast autophagy protein Atg13. epg-1 displays a similar expression pattern to, and directly interacts with, the C. elegans Atg1 homolog UNC-51, suggesting that epg-1 encodes a divergent functional homolog of Atg13 in C. elegans.

The C. elegans ATG101 homolog EPG-9 directly interacts with EPG-1/Atg13 and is essential for autophagy.

Autophagy is an evolutionarily conserved catabolic process that involves the engulfment of cytoplasmic contents in a closed double-membrane structure, called the autophagosome, and their subsequent delivery to the vacuole/lysosomes for degradation. Genetic screens in Saccharomyces cerevisiae have identified more than 30 autophagy-related (Atg) genes that are essential for autophagosome formation. Here we isolated a novel autophagy gene, epg-9, whose loss of function causes defective autophagic degradation of a variety of protein aggregates during C. elegans embryogenesis. Mutations in epg-9 also reduce survival of animals under food depletion conditions. epg-9 mutants exhibit autophagy phenotypes characteristic of those associated with loss of function of unc-51/Atg1 and epg-1/Atg13. epg-9 encodes a protein with significant homology to mammalian ATG101. EPG-9 directly interacts with EPG-1/Atg13. Our study indicates that EPG-9 forms a complex with EPG-1 in the aggrephagy pathway in C. elegans.

Arginine Methylation Modulates Autophagic Degradation of PGL Granules in C. elegans

The selective degradation of intracellular components by autophagy involves sequential interactions of the cargo with a receptor, which also binds the autophagosomal protein Atg8 and a scaffold protein. Here, we demonstrated that mutations in C. elegans epg-11, which encodes an arginine methyltransferase homologous to PRMT1, cause the defective removal of PGL-1 and PGL-3 (cargo)-SEPA-1 (receptor) complexes, known as PGL granules, from somatic cells during embryogenesis. Autophagic degradation of the PGL granule scaffold protein EPG-2 and other protein aggregates was unaffected in epg-11/prmt-1 mutants. Loss of epg-11/prmt-1 activity impairs the association of PGL granules with EPG-2 and LGG-1 puncta. EPG-11/PRMT-1 directly methylates arginines in the RGG domains of PGL-1 and PGL-3. Autophagic removal of PGL proteins is impaired when the methylated arginines are mutated. Our study reveals that posttranslational arginine methylation regulates the association of the cargo-receptor complex with the scaffold protein, providing a mechanism for modulating degradation efficiency in selective autophagy.

The C. elegans engrailed homolog ceh-16 regulates the self-renewal expansion division of stem cell-like seam cells.

Stem cells undergo symmetric and asymmetric division to maintain the dynamic equilibrium of the stem cell pool and also to generate a variety of differentiated cells. The homeostatic mechanism controlling the choice between self-renewal and differentiation of stem cells is poorly understood. We show here that ceh-16, encoding the C. elegans ortholog of the transcription factor Engrailed, controls symmetric and asymmetric division of stem cell-like seam cells. Loss of function of ceh-16 causes certain seam cells, which normally undergo symmetric self-renewal expansion division with both daughters adopting the seam cell fate, to divide asymmetrically with only one daughter retaining the seam cell fate. The human engrailed homolog En2 functionally substitutes the role of ceh-16 in promoting self-renewal expansion division of seam cells. Loss of function of apr-1, encoding the C. elegans homolog of the Wnt signaling component APC, results in transformation of self-renewal maintenance seam cell division to self-renewal expansion division, leading to seam cell hyperplasia. The apr-1 mutation suppresses the seam cell division defect in ceh-16 mutants. Our study reveals that ceh-16 interacts with the Wnt signaling pathway to control the choice between self-renewal expansion and maintenance division and also demonstrates an evolutionarily conserved function of engrailed in promoting cell proliferation.

RNA-binding proteins SOP-2 and SOR-1 form a novel PcG-like complex in C. elegans

We describe the identification and characterization of a novel PcG gene in C. elegans, sor-1, which is involved in global repression of Hox genes. sor-1 encodes a novel protein with an RNA-binding activity. We provide evidence that SOR-1 and the previously identified RNA-binding protein SOP-2 may constitute an RNA-binding complex in Hox gene repression. SOR-1 and SOP-2 directly interact with each other and are colocalized in nuclear bodies. The localization of SOR-1 depends on SOP-2. Surprisingly, homologs of SOR-1 and SOP-2 are not found in other organisms, including the congeneric species C. briggsae, suggesting an unexpected lack of evolutionary constraint on an essential global gene regulatory system.

Selective autophagic degradation of maternally-loaded germline P granule components in somatic cells during C. elegans embryogenesis

Germline P granules are specialized protein/RNA aggregates that are found exclusively in germ cells in C. elegans. During the early embryonic divisions that generate germ blastomeres, aggregate-prone P granule components PGL-1 and PGL-3 that remain in the cytoplasm destined for somatic daughters are selectively removed by autophagy. Loss-of-function of components of the autophagy pathway, including the VPS-34/BEC-1 complex, causes accumulation of PGL-1 and PGL-3 into aggregates in somatic cells (termed PGL granules). Formation of PGL granules depends on SEPA-1, which is an integral component of these granules. SEPA-1 is preferentially degraded by autophagy and is also required for the autophagic degradation of PGL-1 and PGL-3. SEPA-1 functions as a bridging molecule in mediating degradation of P granule components by directly interacting with PGL-3 and also with the autophagy protein LGG-1/Atg8. The defect in embryonic development in autophagy mutants is suppressed by mutation of sepa-1, suggesting that autophagic degradation of PGL granule components may provide nutrients for embryogenesis and/or also prevent the formation of aggregates that could be toxic for animal development. Our study reveals a specific physiological function of selective autophagic degradation during C. elegans development.

Polycomb-like genes are necessary for specification of dopaminergic and serotonergic neurons in Caenorhabditis elegans

The molecular mechanisms underlying the formation of neurons with defined neurotransmitters are not well understood. In this study, we demonstrate that the PcG-like genes in Caenorhabditis elegans, sop-2 and sor-3, regulate the formation of dopaminergic and serotonergic neurons and several other neuronal properties. sor-3 encodes a novel protein containing an MBT repeat, a domain that contains histone-binding activity and is present in PcG proteins SCM and Sfmbt in other organisms. We further show that mutations in sor-3 lead to ectopic expression of Hox genes and cause homeotic transformations. Specification of certain neuronal identities by these PcG-like genes appears to involve regulation of non-Hox gene targets. Our studies revealed that the PcG-like genes are crucial for coordinately regulating the expression of discrete aspects of neuronal identities in C. elegans.