Biology subjects

Liu, J. C.

Publications and source records attributed to Liu, J. C..

8 recordsLinked to original sources

bioRxiv2026

A new tool for quantifying mitochondrial RNA processing intermediates reveals increased levels of immature mRNA species under oxidative stress

Mitochondrial gene expression in mammals requires the processing of polycistronic transcripts into mature RNAs, generating multigene maturation intermediates. Yet these species and their potential association with mitochondrial stress remain poorly characterized. We analyzed intact RNA species in healthy mouse hearts using direct RNA sequencing and identified polyadenylated mitochondrial transcripts containing complete mRNA sequences with uncleaved tRNA extensions, which we termed immature mRNAs (imRNAs). Guided by these findings, we developed a new computational tool to estimate imRNA levels using conventional short-read RNA-seq datasets. Analysis of mouse hearts deficient in mitochondrial superoxide dismutase 2 (SOD2), a model of mitochondrial oxidative stress, revealed selectively elevated levels of imRNAs retaining 5' tRNA extensions, without increases in those with 3' tRNA extensions. Comparative analyses of several mouse models with defects in mitochondrial respiration, transcription, translation, or proteostasis did not reproduce this pattern. In fact, a mouse model of mitochondrial import stress with reduced ROS generation exhibited decreased levels of imRNAs retaining 5' tRNA extensions. Conversely, human induced pluripotent stem cells treated with rotenone, which elevates mitochondrial ROS, showed increased levels of imRNAs retaining 5' but not 3' tRNA extensions, reproducing the pattern observed in SOD2-deficient mouse hearts. These findings identify an imRNA signature potentially associated with mitochondrial oxidative stress. The presence of imRNAs described here suggests that widely used transcriptome analysis pipelines may count these species as mature mRNAs, overestimating mitochondrial mRNA levels and overlooking defects in RNA maturation. By enabling retrospective analysis of processing intermediates, our tool provides access to previously unexamined information in existing RNA-seq datasets.

molecular biology↗

bioRxiv2025

Elucidating cancer cachexia-mediated aberrant cardiac wasting signaling in human iPSC-derived cardiac muscle

Cancer cachexia is a highly debilitating clinical syndrome of involuntary body mass loss featuring profound muscle wasting leading to high mortality. Notably, cardiac wasting is prominent in cancer patients and cancer survivors. Cachexia studies present significant challenges due to the absence of human models and mainly short-term animal studies. To address this translational gap, we have developed a robust human-based cachexia experimental approach characterized by marked cardiac muscle wasting and contractile dysfunction, with increased expression of protein degradation markers. Using human iPSC-derived cardiac muscle, we investigated morphological, functional, and metabolic alterations in the key stages of cachexia and in the post-cachexia phase. C26 and HCT116 tumor cell lines were used to induce cachexia by two methods, pulse addition of cancer cell conditioned media or in transwell-adapted co-culture. Cachectic cardiac myocytes exhibited reduced contraction amplitude, prolonged relaxation time, and increased oxygen consumption rate (OCR), as assessed by video-based and Seahorse analyses. Mechanistic investigations centered on the Atrogin-1/Calcineurin A/NFAT axis revealed this signaling pathway as a central driver of cachexia-induced cardiac atrophy. Cachectic cardiac myocytes exhibited significant upregulation of Atrogin-1, leading to a marked decrease in Calcineurin A protein levels. This, in turn, impaired nuclear translocation of NFAT, thereby suppressing its transcriptional activity and downstream cell growth signaling. These molecular changes were accompanied by increased autophagic flux, as indicated by elevated LC3BII/LC3BI ratios. Furthermore, withdrawal of cachexia-inducing stimuli followed by regular media changes for one week led to normalization of Atrogin-1 and autophagy markers; however, functional impairments and metabolic dysregulation persisted, highlighting delayed recovery. Our new findings establish the Atrogin-1/Calcineurin A/NFAT axis as a key regulatory mechanism in cardiac muscle wasting and suggest this aberrant signaling axis may serve as a targetable mechanism for treatment of cachexia-induced cardiac dysfunction. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=154 HEIGHT=200 SRC="FIGDIR/small/670675v2_ufig1.gif" ALT="Figure 1"> View larger version (31K): org.highwire.dtl.DTLVardef@12e9borg.highwire.dtl.DTLVardef@19ad494org.highwire.dtl.DTLVardef@1fa664corg.highwire.dtl.DTLVardef@5232a6_HPS_FORMAT_FIGEXP M_FIG C_FIG

molecular biology↗

bioRxiv2025

Scalable expansion and hepatic zone maturation of hepatic progenitor cells from human pluripotent stem cells

Scalable generation of functionally mature hepatocytes from human pluripotent stem cells (hPSCs) is vital for regenerative medicine, disease modeling, and drug screening. We present a modular differentiation platform that enables serial expansion and cryopreservation of hepatic progenitors (hepatoblasts; HBs), supporting efficient downstream maturation. By modulating bile acid signaling with FGF19 and culturing under hypoxia, we preserved HB proliferation and bipotency. Subsequent thyroid hormone treatment induced alpha-fetoprotein-negative, albumin-positive hepatocytes, while WNT pathway modulation promoted zonal identity, mimicking in vivo hepatic metabolic heterogeneity. Zonation was confirmed by differential drug-metabolizing enzyme activity, mitochondrial function, and enhanced engraftment in a liver injury mouse model. To evaluate translational relevance, mature hepatocytes were incorporated into extrusion-based 3D bioprinted constructs, which maintained hepatic viability and function. This integrated strategy of combining progenitor cell banking, zonation control, and tissue engineering offers a scalable and clinically relevant approach for generating functional human liver tissue suitable for therapeutic development.

cell biology↗

bioRxiv2025

DNA polymerase beta expression in head & neck cancer modulates the poly(ADP-ribose)-mediated replication checkpoint

Head and Neck Squamous Cell Carcinoma (HNSCC) imposes a significant health burden, necessitating innovative therapeutic strategies to enhance treatment efficacy. Current treatments such as surgery, radiation, and chemotherapy have limited effectiveness and yield severe side effects, emphasizing the need for targeted therapies. We have focused on DNA polymerase beta (Pol{beta}) and its roles in replication stress, cellular responses to DNA damaging therapies, and DNA damage response modifiers. Our investigations reveal a regulatory role for base excision repair (BER) proteins, including Pol{beta}, in the cellular response to inhibitors of poly(ADP-ribose) glycohydrolase (PARG), an enzyme involved in poly(ADP-ribose) (PAR) degradation. The inhibition of PARG, in HNSCC cells, elicits replication stress. Further, this activates the PAR-induced S-phase/ATR checkpoint, leading to a block to replication, cell cycle arrest, and the onset of apoptosis. However, Pol{beta} overexpression mitigates this response, reducing replication-stress induced PAR foci formation, suggesting a modulation of replication checkpoint activation. We found that PARG inhibitor treatment is ineffective on HNSCC cells that overexpress Pol{beta}, implying that the PARG inhibitor-induced PAR and apoptotic response is dependent on the level of Pol{beta}. Further, our in vitro experiments demonstrate that combining PARG and ATR/CHK1 inhibitors overcomes Pol{beta}-mediated treatment resistance in HNSCC cells, producing enhanced effects as compared to the individual treatment conditions. Our findings suggest a possible treatment paradigm for HNSCC, employing ATR or CHK1 inhibitors in combination with PARG inhibitors. This strategy offers a promising path for more effective HNSCC treatments, potentially overcoming Pol{beta}-related resistance.

cancer biology↗

bioRxiv2024

Sox15 is a novel retinal developmental gene that promotes cone photoreceptor differentiation through inhibition of alternative rod photoreceptor fates

The type of cell fate decisions a progenitor can make in the developing nervous system are dependent on a combination of cell intrinsic gene expression programs and cell extrinsic signaling molecules that influence expression programs. Moreover, both the progenitors spatial location in the tissue and time at which it exists during development can, to varying degrees, influence the competency of differentiation. Here, we use the developing mouse retina to understand how signaling factors in the retinal niche can influence a progenitor to become unipotentially-restricted to differentiate into cone photoreceptors, and how this competency changes over the course of retinal development. We show that the secreted inhibitory protein COCO causes progenitors to become cone-restricted, and that the competency for this restricted differentiation is maintained throughout retinal development. Using RNA-sequencing, we identify the transcription factor Sox15 as a potential novel marker of restricted progenitors in the developing retina, and that it acts in response to COCOs effects. Knockdown experiments indicate that this novel retinal transcription factor promotes cone differentiation by inhibiting differentiation into alternative photoreceptor types.

developmental biology↗

bioRxiv2024

Luminal breast epithelial cells from wildtype and BRCA mutation carriers harbor copy number alterations commonly associated with breast cancer

Cancer-associated mutations have been documented in normal tissues, but the prevalence and nature of somatic copy number alterations and their role in tumor initiation and evolution is not well understood. Here, using single cell DNA sequencing, we describe the landscape of CNAs in >42,000 breast epithelial cells from women with normal or high risk of developing breast cancer. Accumulation of individual cells with one or two of a specific subset of CNAs (e.g. 1q gain and 16q, 22q, 7q, and 10q loss) is detectable in almost all breast tissues and, in those from BRCA1 or BRCA2 mutations carriers, occurs prior to loss of heterozygosity (LOH) of the wildtype alleles. These CNAs, which are among the most common associated with ductal carcinoma in situ (DCIS) and malignant breast tumors, are enriched almost exclusively in luminal cells not basal myoepithelial cells. Allele-specific analysis of the enriched CNAs reveals that each allele was independently altered, demonstrating convergent evolution of these CNAs in an individual breast. Tissues from BRCA1 or BRCA2 mutation carriers contain a small percentage of cells with extreme aneuploidy, featuring loss of TP53, LOH of BRCA1 or BRCA2, and multiple breast cancer-associated CNAs in addition to one or more of the common CNAs in 1q, 10q or 16q. Notably, cells with intermediate levels of CNAs are not detected, arguing against a stepwise gradual accumulation of CNAs. Overall, our findings demonstrate that chromosomal alterations in normal breast epithelium partially mirror those of established cancer genomes and are chromosome- and cell lineage-specific.

cancer biology↗

bioRxiv2023

Sea squirt-inspired bio-derived tissue sealants

Sea squirts or tunicates bodies are composed of cellulose nanofibers and gallol- functionalized proteins. These sea creatures are known to heal their injuries under seawater by forming crosslinks between gallols and functional groups from other proteins in their bodies. Inspired by their wound healing mechanism, herein, we have developed a tissue sealant using zein (a plant-based protein) and tannic acid (gallol-containing polyphenol). Except for fibrin- based sealants, most commercial surgical adhesives, and sealants available today are derived from petroleum products that compromise their biodegradability. They often have complicated and multi-step synthesis processes that ultimately affect their affordability. To overcome this challenge, we ensured that these sea squirt-inspired tissue sealants are bio-based, easily synthesized, and low-cost. The sealants were studied on their own and with a food-grade enzyme transglutaminase. The adhesion performances of the sealants were found to be higher than physiological pressures in seven out of nine different tissue substrates studied here. Their performance was also better than or on par with the FDA-approved fibrin sealant Tisseel. Ex vivo models demonstrate instant sealing of leaking wounds in less than a minute. The sealants were not only cytocompatible but also showed complete wound healing on par with sutures and Tisseel when applied in vivo on skin incisions in rats. Overall, these sea squirt-inspired bio-based sealants show great potential to replace currently available wound closure methods.

bioengineering↗

bioRxiv2021

Single Cell, Single Nucleus and Spatial RNA Sequencing of the Human Liver Identifies Hepatic Stellate Cell and Cholangiocyte Heterogeneity

The critical functions of the human liver are coordinated through the interactions of hepatic parenchymal and non-parenchymal cells. Recent advances in single cell transcriptional approaches have enabled an examination of the human liver with unprecedented resolution. However, dissociation related cell perturbation can limit the ability to fully capture the human livers parenchymal cell fraction, which limits the ability to comprehensively profile this organ. Here, we report the transcriptional landscape of 73,295 cells from the human liver using matched single-cell RNA sequencing (scRNA-seq) and single-nucleus RNA sequencing (snRNA-seq). The addition of snRNA-seq enabled the characterization of interzonal hepatocytes at single-cell resolution, revealed the presence of rare subtypes of hepatic stellate cells previously only seen in disease, and detection of cholangiocyte progenitors that had only been observed during in vitro differentiation experiments. However, T and B lymphocytes and NK cells were only distinguishable using scRNA-seq, highlighting the importance of applying both technologies to obtain a complete map of tissue-resident cell-types. We validated the distinct spatial distribution of the hepatocyte, cholangiocyte and stellate cell populations by an independent spatial transcriptomics dataset and immunohistochemistry. Our study provides a systematic comparison of the transcriptomes captured by scRNA-seq and snRNA-seq and delivers a high-resolution map of the parenchymal cell populations in the healthy human liver.

cell biology↗