Biology subjects

Liu, K. H.

Publications and source records attributed to Liu, K. H..

4 recordsLinked to original sources

bioRxiv2026

Inducible Impairment of Polymerase Gamma Activity in Cardiomyocytes Promotes Severe Cardiomyopathy with Cardiac Hepatopathy

Mitochondrial dysfunction is a hallmark feature of heart failure (HF) and cardiomyopathy, with substantial human and preclinical evidence suggesting that congenital mitochondrial defects can directly drive these conditions. Despite these strong links, not all individuals with mitochondrial disease develop cardiomyopathy, and therefore the precise mitochondrial defects that initiate cardiac pathology in this setting remain incompletely understood. Here, we describe a novel mouse model that induces progressive deterioration in mtDNA integrity specifically in cardiomyocytes. This model was generated through a post developmental, cardiomyocyte specific deletion of the exonuclease domain of Polymerase Gamma (PolG), impairing its ability to repair mtDNA. Strikingly, from just 16 weeks post-induction these mice displayed progressive worsening of cardiac output, ejection fraction, global strain and blood pressure culminating in premature death at 28-30 weeks post-mutation. Morphologically, these mice displayed cardinal features of hypertrophic cardiomyopathy with enlarged hearts, ventricles and cardiac fibrosis - but little evidence of congestive HF. We also demonstrate using various transcriptional and proteomic readouts, that signalling pathways characteristic of cardiomyopathy were activated. Interestingly, prior to substantial declines in cardiac function, we detect robust activation of the integrated stress response (ISR) in PolGMut mice, and major rewiring of mitochondrial folate metabolism pathways, suggesting that these pathways underpin the developing pathology. Lastly, we also observed a striking hepatopathy phenotype in mutant mice reminiscent of that observed in patients with HF, a condition not robustly recapitulated previously in other mouse models. Thus, our data establish a direct link between mtDNA instability leading to chronic activation of the ISR and rewiring of folate metabolism in cardiomyocytes, subsequently driving a severe cardiac phenotype associated with hepatopathy.

physiology↗

bioRxiv2022

High-throughput Production of Diverse Xenobiotic Metabolites with P450-transduced Huh7 Hepatoma Cell Lines

Precision medicine requires methods to assess drug metabolism and distribution, including the identification of known and undocumented drug and chemical exposures as well as their metabolites. Recent work demonstrated high-throughput generation of xenobiotic metabolites with human liver S-9 fractions and detection in human plasma and urine. Here, we developed a panel of lentivirally transduced human hepatoma cell lines (Huh7) that stably express individual cytochrome P450 (P450) enzymes and generate P450-specific xenobiotic metabolites. We verified protein expression by immunoblotting and demonstrated that the cell lines generate P450-specific metabolites from probe substrates. To increase analytical throughput, we used a pooling strategy where 36 chemicals were grouped into 12 unique mixtures, each mixture containing 6 randomly selected compounds, and each compound being present in two separate mixtures. Each mixture of compounds was incubated with 8 different P450 cell lines with cell extracts analyzed at 0 and 2 h. Extracts were analyzed using liquid chromatography-high resolution mass spectrometry. Cell lines selectively metabolized test substrates, with pazopanib metabolized by CYP3A4 and CYP2C8 cells, bupropion by CYP2B6, and {beta}-naphthoflavone by CYP1A2 for example, showing substrate-enzyme specificity. Predicted metabolites from the remaining 33 compounds as well as many unidentified m/z features were detected. We also show that a specific metabolite generated by CYP2B6 cells, but not detected in the S9 system, was identified in human samples. Our data show that incubating these cell lines with chemical mixtures accelerated characterization of xenobiotic chemical space, while simultaneously allowing for the contributions of specific P450 enzymes to be identified. Significance statementHigh resolution mass spectrometry enables the identification of exposures to drugs and other xenobiotics in human samples. This paper demonstrates a workflow for high throughput production of xenobiotic metabolites using a panel of engineered cytochrome P450-expressing hepatoma cells. Active substrate-enzyme pairs can be identified using this workflow and generated metabolites can be used as surrogate standards to validate xenobiotic detection in humans.

pharmacology and toxicology↗

bioRxiv2021

α1-COP delivers sphingolipid modifiers and controls plasmodesmal callose deposition in Arabidopsis

Callose is a plant cell wall polymer in the form of {beta}-1,3-glucan, which regulates symplasmic channel size at plasmodesmata (PD). It plays a crucial role in a variety of processes in plants through the regulation of intercelluar symplasmic continuity. However, how to maintain callose homeostasis at PD in the molecular levels is poorly understood. To further elucidate the mechanism of PD callose homeostasis, we screened and identified an Arabidopsis mutant plant that exhibited excessive callose deposition at PD. Based on the Next-generation sequencing (NGS)-based mapping, other mutant allele analysis, and complementation assay, the mutated gene was shown to be 1-COP, which encodes a member of the COPI coatomer complex comprised of , {beta}, {beta}', {gamma}, {delta}, {varepsilon}, and {zeta} subunits. Since there is no report on the link between COPI and callose/PD, it was extremely curious to know the roles of 1-COP or COPI in PD regulation through callose deposition. Here, we report that loss-of-function of 1-COP directly elevates the callose accumulation at PD by affecting subcellular protein localization of callose degradation enzyme PdBG2. This process is linked to ERH1, an inositol phosphoryl ceramide synthase (IPCS), and glucosylceramide synthase (GCS) functions through physical interactions with the 1-COP protein. In addition, the loss-of-function of 1-COP also alters the subcellular localization of ERH1 and GCS proteins, results in a reduction of GlcCers and GlcHCers molecules, which are the key SL species for lipid raft formation. According to our findings, we propose that 1-COP protein, together with the SL modifiers controlling lipid raft compositions, regulates the function of GPI-anchored PD proteins and hence the callose turnover at PD and symplastic movement of biomolecules. Our findings provide the first key clue to link the COPI-mediated intracellular trafficking pathway to the callose-mediated intercellular signaling pathway through PD. One-sentence summaryPlant-specific coatomer protein functions as a negative regulator of callose accumulation by regulating the translocation of sphingolipid enzymes.

cell biology↗

bioRxiv2020

Glucosylhydroxyceramides Modulate Secretion Machinery of a Subset of Plasmodesmata Proteins and a Change in the Callose Accumulation

The plasma membranes encapsulated in the plasmodesmata (PDs) with symplasmic nano-channels contain abundant lipid rafts, which are enriched by sphingolipids and sterols. The attenuation of sterol compositions has demonstrated the role played by lipid raft integrity in the intercellular trafficking of glycosylphosphatidylinositol (GPI)-anchored PD proteins, particularly affecting in the callose enhancement. The presence of callose at PD is tightly attributed to the callose metabolic enzymes, callose synthases (CalSs) and {beta}-1,3-glucanases (BGs) in regulating callose accumulation and callose degradation, respectively. Sphingolipids have been implicated in signaling and membrane protein trafficking, however the underlying processes linking sphingolipid compositions to the control of symplasmic apertures remain unknown. A wide variety of sphingolipids in plants prompts us to investigate which sphingolipid molecules are important in regulating symplasmic apertures. Here, we demonstrate that perturbations of sphingolipid metabolism by introducing several potential sphingolipid (SL) pathway inhibitors and genetically modifying SL contents from two independent SL pathway mutants are able to modulate callose deposition to control symplasmic connectivity. Our data from pharmacological and genetic approaches show that the alteration in glucosylhydroxyceramides (GlcHCers) particularly disturb the secretory machinery for GPI-anchored PdBG2 protein, resulting in an over accumulated callose. Moreover, our results reveal that SL-enriched lipid rafts link symplasmic channeling to PD callose homeostasis by controlling the targeting of GPI-anchored PdBG2. This study elevates our understanding of the molecular linkage underlying intracellular trafficking and precise targeting to specific destination of GPI-anchored PD proteins incorporated with GlcHCers contents.

plant biology↗