Biology subjects

Chi, Q.

Publications and source records attributed to Chi, Q..

11 recordsLinked to original sources

bioRxiv2025

WormTagDB: A Systematic Survey of Endogenously Tagged Proteins in C. elegans and Roadmap Towards the Tagged Proteome

Endogenous protein tagging in Caenorhabditis elegans enables direct visualization and manipulation of proteins in vivo, providing native readouts of expression, localization, and dynamics. No coordinated effort currently exists to comprehensively tag proteins on a large scale, resulting in patchy coverage that limits comprehensive proteome analyses. We systematically reviewed 2,500 primary research articles, identifying 778 that report novel endogenous tags, and integrated these with the Caenorhabditis Genetics Center strain records to catalog >90% of all existing tagged alleles. In total, we found that 1,554 unique genes (~8% of the proteome) have been endogenously tagged. Gene Ontology enrichment analysis revealed that cytoskeletal proteins, transcription factors, and RNA-binding proteins dominate the tagged proteome, while membrane proteins, metabolic enzymes, and mitochondrial components remain largely untagged, reflecting both technical barriers and research priorities that have shaped the last decade of tagging efforts. We created WormTagDB (https://wormtagdb.rc.duke.edu), an interactive, community-updatable resource that consolidates all known endogenously tagged alleles and provides precomputed CRISPR guide and homology-arm primer designs for N- and C-terminal knock-ins across all protein-coding genes. This will enable researchers to easily identify existing alleles to prevent redundant strain generation and rapidly initiate new knock-in experiments. A systematic effort to tag every C. elegans gene would deliver the first complete metazoan visual proteome, providing comprehensive insights into protein localization, dynamics, and regulation, revealing new protein associations and molecular processes.

genetics↗

bioRxiv2025

Specialized high-capacity mitochondria fuel cell invasion

Cell invasion through basement membrane (BM) is energetically intensive, and how an invading cell produces high ATP levels to power invasion is understudied. By generating 20 endogenously tagged mitochondrial proteins, we identified a specialized mitochondrial subpopulation within the C. elegans anchor cell (AC) that localizes to the BM breaching site and generates elevated ATP to fuel invasion. These ETC-enriched high-capacity mitochondria are compositionally unique, harboring increased protein import machinery and dense cristae enriched with ETC components. High-capacity mitochondria emerge at the time of AC specification and depend on the AC pro-invasive transcriptional program. Finally, we show that netrin signaling through a Src kinase directs microtubule polarization, which facilitates metaxin adaptor complex dependent ETC-enriched mitochondrial trafficking to the AC invasive front. Our studies reveal that an invasive cell produces high ATP by generating and localizing high-capacity mitochondria. This might be common strategy used by other cells to meet energy demanding processes.

cell biology↗

bioRxiv2025

Mitophagy at the oocyte-to-zygote transition promotes species immortality

The quality of inherited mitochondria determines embryonic viability1, metabolic health during adulthood and future generation endurance. The oocyte is the source of all zygotic mitochondria2, and mitochondrial health is under strict developmental regulation during early oogenesis3-5. Yet, fully developed oocytes exhibit the presence of deleterious mitochondrial DNA (mtDNA)6,7 and mitochondrial dysfunction from high levels of endogenous reactive oxygen species8 and exogenous toxicants9. How fully developed oocytes prevent transmission of damaged mitochondria to the zygotes is unknown. Here we discover that the onset of oocyte-to-zygote transition (OZT) developmentally triggers a robust and rapid mitophagy event that we term mitophagy at OZT (MOZT). We show that MOZT requires mitochondrial fragmentation, activation of the macroautophagy system and the mitophagy receptor FUNDC1, but not the prevalent mitophagy factors PINK1 and BNIP3. Oocytes upregulate expression of FUNDC1 in response to diverse mitochondrial insults, including mtDNA mutations and damage, uncoupling stress, and mitochondrial dysfunction, thereby promoting selection against damaged mitochondria. Loss of MOZT leads to increased inheritance of deleterious mtDNA and impaired bioenergetic health in the progeny, resulting in diminished embryonic viability and the extinction of descendent populations. Our findings reveal FUNDC1-mediated MOZT as a mechanism that preserves mitochondrial health during the mother-to-offspring transmission and promotes species continuity. These results may explain how mature oocytes from many species harboring mutant mtDNA give rise to healthy embryos with reduced deleterious mtDNA.

cell biology↗

bioRxiv2025

On-demand delivery of fibulin-1 enables repeated basement membrane stretching

Basement membrane (BM) extracellular matrices enwrap and structurally support tissues. Whether BMs are uniquely constructed to support tissues that undergo repetitive stretching and recoil events is unknown. During C. elegans ovulation, the spermathecal BM stretches [~]1.7-fold and then returns to its original shape every twenty minutes to passage hundreds of oocytes. Through live fluorescence microscopy, we discovered that ovulating oocytes secrete and deliver the fibulin-1 extracellular matrix protein to the spermathecal BM during stretching, where it forms a dynamic overlapping network with type IV collagen. Fibulin-1 depletion led to a breakdown in type IV collagen and BM organization, resulting in a more deformable BM and extended spermatheca. Moreover, perturbation to fibulin-1 network formation via mutagenesis was sufficient to disrupt organ shape. Together, our study reveals an on-demand fibulin-1 delivery system that protects the BM network when it is stretched, thereby allowing repeated rounds of organ expansion and recovery.

cell biology↗

bioRxiv2024

A complete collagen IV fluorophore knock-in toolkit reveals -chain diversity in basement membrane

The type IV collagen triple helix, composed of three -chains, is a core basement membrane (BM) component that assembles into a network within BMs. Endogenous tagging of all -chains with genetically encoded fluorophores has remained elusive, limiting our understanding of this crucial BM component. Through genome editing, we show that the C-termini of the C. elegans type IV collagen -chains EMB-9 and LET-2 can be fused to a variety of fluorophores to create a strain toolkit with wild-type health. Using quantitative imaging, our results suggest a preference for LET-2-LET-2-EMB-9 trimer construction, but also tissue-specific flexibility in trimers assembled driven by differences in -chain expression levels. By tagging emb-9 and let-2 mutants that model human Gould Syndrome, a complex multi-tissue disorder, we further discover defects in extracellular accumulation and turnover that might help explain disease pathology. Together, our findings identify a permissive tagging site that will allow diverse studies on type IV collagen regulation and function in animals. SummarySrinivasan et al., construct a collagen IV fluorophore knock-in toolkit in C. elegans using a newly identified permissive genome editing site and reveal tissue-specific -chain diversity and basement membrane turnover defects in collagen IV mutants modeling human COL4A1/A2 (Gould) syndrome.

developmental biology↗

bioRxiv2024

De novo lipid synthesis and polarized prenylation drives cell invasion through basement membrane

SummaryInvasive cells form large, specialized protrusions to break through basement membrane (BM) matrix barriers. Park et al., reveal a crucial requirement for de novo lipid synthesis and a dynamic polarizing prenylation system to rapidly construct invasive protrusions that breach BMs. To breach basement membrane (BM), cells in development and cancer use large, transient, specialized lipid-rich membrane protrusions. Using live imaging, endogenous protein tagging, and cell-specific RNAi during C. elegans anchor cell (AC) invasion, we demonstrate that the lipogenic SREBP transcription factor SBP-1 drives expression of the fatty acid synthesis enzymes POD-2 and FASN-1 prior to invasion. We show that phospholipid producing LPIN-1 and sphingomyelin synthase SMS-1, which use fatty acids as substrates, produce lysosome stores that build the ACs invasive protrusion, and that SMS-1 also promotes protrusion localization of the lipid raft partitioning ZMP-1 matrix metalloproteinase. Finally, we discover that the endoplasmic reticulum (ER)-associated HMG-CoA reductase HMGR-1, which generates isoprenoids for prenylation, enriches at the AC invasive front, and that the final ER prenylation enzyme, ICMT-1, localizes to ER exit sites that dynamically polarize to deliver prenylated GTPases for protrusion formation. Together, these results reveal a collaboration between lipogenesis and a polarized lipid prenylation system that drives invasive protrusion formation.

cell biology↗

bioRxiv2023

Reciprocal discoidin domain receptor signaling strengthens integrin adhesion to connect adjacent tissues

Separate tissues connect through adjoining basement membranes to carry out molecular barrier, exchange, and organ support functions. Cell adhesion at these connections must be robust and balanced to withstand independent tissue movement. Yet, how cells achieve synchronized adhesion to connect tissues is unknown. Here, we have investigated this question using the C. elegans utse-seam tissue connection that supports the uterus during egg-laying. Through genetics, quantitative fluorescence, and cell specific molecular disruption, we show that type IV collagen, which fastens the linkage, also activates the collagen receptor discoidin domain receptor 2 (DDR-2) in both the utse and seam. RNAi depletion, genome editing, and photobleaching experiments revealed that DDR-2 signals through LET-60/Ras to coordinately strengthen an integrin adhesion in the utse and seam that stabilizes their connection. These results uncover a synchronizing mechanism for robust adhesion during tissue connection, where collagen both affixes the linkage and signals to both tissues to bolster their adhesion.

developmental biology↗

bioRxiv2022

The C. elegans Anchor Cell Transcriptome: Ribosome Biogenesis Drives Cell Invasion through Basement Membrane

Cell invasion through basement membrane (BM) barriers is important in development, immune function, and cancer progression. As invasion through BM is often stochastic, capturing gene expression profiles of cells actively transmigrating BM in vivo remains elusive. Using the stereotyped timing of C. elegans anchor cell (AC) invasion, we generated an AC transcriptome during BM breaching. Through a focused RNAi screen of transcriptionally enriched genes, we identified new invasion regulators, including TCTP (Translationally Controlled Tumor Protein). We also discovered gene enrichment of ribosomal proteins. AC-specific RNAi, endogenous ribosome labeling, and ribosome biogenesis analysis revealed a burst of ribosome production occurs shortly after AC specification, which drives the translation of proteins mediating BM removal. Ribosomes also strongly localize to the ACs endoplasmic reticulum (ER) and the endomembrane system expands prior to invasion. We show that AC invasion is sensitive to ER stress, indicating a heightened requirement for translation of ER trafficked proteins. These studies reveal key roles for ribosome biogenesis and endomembrane expansion in cell invasion through BM and establish the AC transcriptome as a resource to identify mechanisms underlying BM transmigration.

cell biology↗

bioRxiv2021

Hemicentin mediated type IV collagen assembly strengthens juxtaposed basement membrane linkage

Basement membrane (BM) matrices surround and separate most tissues. However, through poorly understood mechanisms, BMs of adjacent tissues can also stably link to support organ structure and function. Using endogenous knock-in fluorescent proteins, conditional RNAi, optogenetics, and quantitative live imaging, we identified matrix proteins mediating a BM linkage (B-LINK) between the uterine utse and epidermal seam cell BMs in Caenorhabditis elegans that supports the uterus during egg-laying. We found that hemicentin is secreted by the utse and promotes fibulin-1 assembly to jointly initiate the B-LINK. During egg-laying, however, both proteins decline in levels and are not required for B-LINK maintenance. Instead, we discovered that hemicentin also promotes type IV collagen assembly, which accumulates to high levels during egg-laying and sustains the B-LINK during the mechanically active egg-laying period. This work reveals mechanisms underlying BM-BM connection maturation and identifies a crucial function for hemicentin and fibulin-1 in initiating attachment and type IV collagen in strengthening this specialized form of tissue linkage. SummaryTissue attachment through linking juxtaposed basement membranes (BMs) is crucial for the structure and function of many organs. Gianakas et al. identify a key role for hemicentin and fibulin-1 in initiating BM-BM attachment and type IV collagen in stabilizing this linkage and allowing it to resist high mechanical loads.

developmental biology↗

bioRxiv2021

A basement membrane discovery pipeline uncovers network complexity, new regulators, and human disease associations

Basement membranes (BMs) are ubiquitous extracellular matrices whose composition remains elusive, limiting our understanding of BM regulation and function. By developing a bioinformatic and in vivo discovery pipeline, we define a network of 222 human proteins localized to BMs. Network analysis and screening in C. elegans and zebrafish identified new BM regulators, including ADAMTS, ROBO, and TGF{beta}. More than 100 BM-network genes associate with human phenotypes and by screening 63,039 genomes from families with rare disorders, we discovered loss-of-function variants in LAMA5, MPZL2, and MATN2, and show they regulate BM composition and function. This cross-disciplinary study establishes the immense complexity and role of BMs in human health.

cell biology↗

bioRxiv2019

α-integrins dictate distinct modes of type IV collagen recruitment to basement membranes

Basement membranes (BMs) are cell-associated extracellular matrices that support tissue integrity, signaling, and barrier properties. Type IV collagen is critical for BM mechanical and signaling functions, yet how it is directed into BMs in vivo is unclear. Through live-cell imaging of endogenous localization, conditional knockdown, and misexpression experiments, we uncovered distinct mechanisms of integrin-mediated collagen recruitment to Caenorhabditis elegans gonadal and pharyngeal BMs. The putative laminin-binding INA-1/{beta}PAT-3 integrin was selectively activated in the gonad and recruited laminin, which directed moderate collagen incorporation. In contrast, the putative Arg-Gly-Asp (RGD)-binding PAT-2/{beta}PAT-3 integrin was activated in the pharynx and recruited high levels of collagen independent of laminin. Through an RNAi screen, we further identified the small GTPase RAP-3 (Rap1) as a pharyngeal-specific PAT-2/PAT-3 activator that modulates collagen levels. Together, these studies demonstrate that tissues can use distinct mechanisms to direct collagen incorporation into BMs to precisely control collagen levels and construct diverse BMs.

cell biology↗