Biology subjects

Huo, H.

Publications and source records attributed to Huo, H..

10 recordsLinked to original sources

bioRxiv2026

A J-protein-licensed CHD3 chromatin remodeling complex promotes H3K27me3 spreading to maintain cell identity in plants

Stable inheritance of epigenetic states is essential for cellular identity and developmental robustness and requires ATP-dependent chromatin remodeling, yet the molecular composition and regulatory mechanisms of plant CHD3 chromatin remodelers have remained unknown. Here we show that the Arabidopsis CHD3 remodeler PICKLE (PKL) assembles with a defined set of J-domain proteins (JDPs) to form stable CHD3 chromatin remodeling complexes in plants that are compositionally distinct from their animal counterparts. Loss of JDPs phenocopies pkl mutants, and JDPs are required for nucleosome densification and H3K27me3 spreading at thousands of Polycomb target genes. Mechanistically, JDPs perform dual functions by promoting PKL protein stability while directly stimulating PKL chromatin remodeling activity through their plant-specific DUF3444 domain that binds the N-terminal tail of histone H3. Deletion of the DUF3444 domain severely impairs PKL remodeling activity without disrupting complex assembly, leading to defective H3K27me3 spreading, impaired facultative heterochromatin maintenance, and loss of cell identity. Together, our findings define plant CHD3 chromatin remodeling complexes and reveal a mechanism that regulates CHD3-mediated chromatin remodeling to support the propagation of repressive chromatin states in plants.

plant biology↗

bioRxiv2026

An indirect organogenesis-based citrus transformation system monitored using visible reporter markers

Genetic transformation of citrus is essential for functional genomics and trait improvement, yet remains limited by prolonged regeneration cycles, genotype-dependent responses, and inefficient screening of transformed tissues. Here, we established an Agrobacterium-mediated indirect organogenesis-based transformation system for Carrizo citrange and evaluated two visible reporter systems, the anthocyanin regulator ROSEA1 and the betalain biosynthetic cassette RUBY, for monitoring transformed tissues throughout regeneration. An optimized regeneration protocol enabled visible callus ini-tiation within 7 days following a 5-day pre-culture treatment and recovery of PCR-confirmed transgenic plantlets within 5 months. Both visible reporters were compatible with callus development and shoot regeneration throughout the indirect organogenesis workflow. During callus proliferation, visible pigmentation closely corresponded with GFP fluorescence, enabling rapid, non-destructive identification of transformed tissues. However, reporter performance diverged during shoot regeneration: RUBY maintained stable pigmentation throughout regeneration, whereas ROSEA1-associated pigmentation progressively declined despite continued GFP fluorescence. Consequently, RUBY exhibited a 4.7-fold higher pigmenta-tion-based detection rate than ROSEA1 at the shoot stage and showed closer agreement with GFP-based detection. Putative transgenic events were confirmed by PCR in independent lines of both constructs, together with GFP fluorescence in leaves and root tips. Together, these findings establish an efficient indirect organogenesis-based transformation platform for citrus and demonstrate that both ROSEA1 and RUBY are effective visual reporters during callus proliferation, whereas RUBY provides more reliable visual identification during shoot regeneration and plant recovery.

plant biology↗

bioRxiv2026

Whole-body Super-resolution Functional and Molecular Imaging with Panoramic Photoacoustic-Ultrasound Tomography

Photoacoustic (PA) and ultrasound (US) imaging provide complementary molecular, functional, and anatomical contrasts. Here, we present a panoramic PA-US imaging platform that integrates multispectral PA computed tomography (PACT) along with reflection-mode and transmission-mode US imaging through a single shared full-ring ultrasound array. We employ an ultrafast planewave transmission scheme in reflection-mode US for power Doppler (PWD) imaging and ultrasound localization microscopy (ULM). Additionally, we use the transmission-mode US to reconstruct a spatially resolved speed of sound (SoS) map that corrects both PA and US reconstruction. Such correction sharpens the resolution of PACT, suppresses the artifacts of PWD, and improves microbubble localization of ULM. Elevational scanning further enables whole-body volumetric imaging with co-registered PA and US contrasts. The integrated system maps photoswitchable DrBphP1-expressing tumors alongside their blood perfusion and oxygenation environment. Applying the platform to monitor unilateral renal ischemia-reperfusion injury, we report that microvascular perfusion and renal oxygenation recover at different rates. Collectively, we demonstrate that the integrated PA-US imaging platform provides a unified framework for multiparametric study of anatomy, perfusion, microvascular flow, oxygenation, and molecular activities.

bioengineering↗

bioRxiv2026

miR319-targeted LsTCP4 and non-target LsTCP17 act in parallel to promote leaf senescence in lettuce

Leaf senescence directly affects lettuce quality and postharvest shelf life, but the regulatory roles of miR319-targeted and non-target CIN-TCP transcription factors remain unclear. Here, we examined whether the miR319-TCP module controls lettuce leaf senescence through separable genetic branches. MIR319 overexpression delayed dark-induced senescence, whereas STTM-mediated miR319 suppression accelerated chlorophyll loss, photosynthetic decline, and senescence-marker activation. Disruption of the miR319-targeted gene LsTCP4 phenocopied MIR319 overexpression, supporting LsTCP4 as a pro-senescence factor downstream of miR319. We further found that the miR319 non-target CIN gene LsTCP17 also promoted senescence, as tcp17 leaves retained more chlorophyll than wild type during dark treatment. Genetic combinations showed that tcp17 enhanced chlorophyll retention in the OX319 background and partially rescued the accelerated senescence phenotype of S319, indicating that LsTCP17 acts through a route separable from the miR319-targeted branch. Together, these results reveal a split CIN-TCP architecture in which miR319-targeted LsTCP4 and non-target LsTCP17 provide parallel pro-senescence inputs, offering a genetic framework for targeted improvement of lettuce quality.

plant biology↗

bioRxiv2026

miR319 promotes de novo shoot regeneration by repressing LsTCP4 in lettuce

Plant regeneration is a major determinant of transformation and genome-editing efficiency, yet the endogenous regulatory networks controlling regenerative competence in horticultural crops remain incompletely understood. The miR319-TCP module regulates multiple developmental processes in plants, but its function in lettuce regeneration has not been defined. Here, we performed a genome-wide analysis of the TEOSINTE BRANCHED1/CYCLOIDEA/PROLIFERATING CELL FACTOR (TCP) gene family in lettuce (Lactuca sativa). Thirty-three LsTCP genes were identified and classified into Class I/PCF, Class II/CIN, and Class II/CYC/TB1 groups. Five CIN-class genes, LsTCP2, LsTCP3, LsTCP4, LsTCP10, and LsTCP24, were predicted as high-confidence miR319 targets and supported by degradome-based cleavage evidence. MIR319-overexpression (OX319) explants showed enhanced de novo shoot regeneration, with 94.5% regeneration efficiency and 1.92 shoots per explant, whereas STTM-miR319 suppression (S319) explants showed reduced regeneration, with 28.5% regeneration efficiency and 0.36 shoots per explant. These phenotypes were associated with altered expression of several miR319-targeted CIN-TCP genes, particularly LsTCP4, LsTCP10, and LsTCP24. Disruption of LsTCP4 increased regeneration efficiency to 91.4% and shoot production to 2.05 shoots per explant, resembling the regeneration-enhancing effect of miR319 overexpression. In contrast, disruption of the non-target CIN gene LsTCP17 did not significantly affect regeneration under the tested conditions. Together, these results identify LsTCP4 as a key miR319-responsive negative regulator of de novo shoot regeneration and highlight miR319-mediated repression of LsTCP4 as a potential endogenous strategy for improving lettuce regeneration.

plant biology↗

bioRxiv2026

3D Passive Cavitation Mapping (3D-PCM) with a Large-aperture Planar Array

Urinary stone disease is a common urological condition with increasing incidence, particularly in developed countries. Laser lithotripsy (LL) has become a preferred minimally invasive treatment due to its high precision and low tissue damage. Recent studies suggest that cavitation plays a critical role in stone damage during LL, and three-dimensional passive cavitation mapping (3D-PCM) has emerged as a promising tool for detecting these events. However, clinical translation of 3D-PCM remains challenging due to limitations in imaging depth, field of view (FOV), and procedural compatibility. Here, we present a large-FOV dual-modality imaging system (3D-PCM and B-mode ultrasound) based on a large-aperture planar ultrasound array. Through array optimization and model-based reconstruction, our system achieves an expanded FOV of [~]40 x 40 mm2 at a clinically relevant imaging depth of [~]110 mm, while maintaining high spatial resolution of [~]0.6 mm laterally and [~]0.4 mm axially. In vivo experiments in a porcine model demonstrate that the reconstructed cavitation distribution correlates well with stone damage. Our technology has the potential to provide real-time treatment feedback during LL without disrupting the standard workflow.

bioengineering↗

bioRxiv2026

Three-Dimensional Photoacoustic Tomography with Ultrasound Localization Priors

Three-dimensional photoacoustic tomography (3D-PAT) enables noninvasive structural and functional imaging with optical absorption contrast and ultrasonic detection depth. However, its spatial resolution is limited by acoustic diffraction, and incomplete detection geometry can substantially degrade image fidelity and quantitative accuracy. Here, we present a ULM-guided model-based reconstruction framework, termed 3D-PAULMprior that incorporates sub-diffraction vascular priors from concurrent ultrasound localization microscopy (ULM) into 3D photoacoustic reconstruction. The method uses weighted regional Laplacian regularization to integrate high-resolution vascular information into the inverse problem, thereby enhancing vascular sharpness, suppressing limited-view artifacts, and improving blood oxygen saturation estimation. We validated 3D-PAULMprior using numerical simulations, tissue-mimicking phantoms, and in vivo mouse brain imaging. Compared with conventional reconstruction, 3D- PAULMprior improved spatial resolution by over 50%, increased contrast-to-noise ratio by 261.2%, and enhanced structural similarity index by 24.6%. In vivo, 3D-PAULMprior recovered vascular structures that were poorly resolved or missing in conventional reconstructions and produced more spatially confined sO2 maps. These results establish 3D-PAULMprior as a robust multimodal reconstruction strategy for high-resolution structural and functional photoacoustic imaging.

bioengineering↗

bioRxiv2023

Optimization of Shoot Regeneration and Application of CRISPR/Cas9 Gene Editing to Cultivated Strawberry

Efficient methods of plant transformation and tissue culture are essential to CRISPR/Cas-based gene editing of crops, but neither is well established in cultivated octoploid strawberry (F xananassa). In the present study, a method for shoot regeneration was established and optimized for two strawberry cultivars commercially grown in Florida, Sweet Sensation(R) Florida 127 FL127) and Florida Brilliance (FB). Runner segments at the tip, node, and petiole obtained from greenhouse-grown plants were used as explants for comparisons of shoot regeneration rate. FL127 showed the highest frequency of shoot regeneration to a basal Murashige and Skoog media (MS) containing 1 mg{middle dot}L-1 of TDZ, 0.05 mg{middle dot}L-1 of BA, and 0.05 mg{middle dot}L-1 of 2,4-D, while FB showed the best response to a lower concentration of BA (0.01 mg{middle dot}L-1) in the same media type. The average conversion frequencies of somatic embryos into shoot regenerations from the runner tips (RT) were 42.8% in FL127 and 56.9% in FB, respectively, with RT being the most prolific in shoot generation for both cultivars. Using these optimized tissue conditions, Agrobacterium-mediated CRISPR/Cas9 gene editing was conducted to evaluate the efficiency of transformation and knockout mutations in the phytoene desaturase (FaPDS) gene of FL127. A total of 234 explants treated with Agrobacterium resulted in an 80.3% regeneration efficiency, with 13.3% of regenerated plants exhibiting partial or complete albino phenotypes. Amplicon sequencing of edited progeny revealed substitutions, insertions, and deletions at the gRNA target sites or flanking regions of all FaPDS homoeologous copies. Our results provide effective methods of tissue culture and transformation for the efficient application of CRISPR-mediated gene editing in cultivated strawberry.

plant biology↗

bioRxiv2022

Development of highly potent non-covalent inhibitors of SARS-CoV-2 3CLpro

The SARS-CoV-2 virus is the causal agent of the ongoing pandemic of coronavirus disease 2019 (COVID-19). There is an urgent need for potent, specific antiviral compounds against SARS-CoV-2. The 3C-like protease (3CLpro) is an essential enzyme for the replication of SARS-CoV-2 and other coronaviruses, and thus is a target for coronavirus drug discovery. Nearly all inhibitors of coronavirus 3CLpro reported so far are covalent inhibitors. Here, we report the development of specific, non-covalent inhibitors of 3CLpro. The most potent one, WU-04, effectively blocks SARS-CoV-2 replications in human cells with EC50 values in the 10-nM range. WU-04 also inhibits the 3CLpro of SARS-CoV and MERS-CoV with high potency, indicating that it is a pan-inhibitor of coronavirus 3CLpro. WU-04 showed anti-SARS-CoV-2 activity similar to that of PF-07321332 (Nirmatrelvir) in K18-hACE2 mice when the same dose was administered orally. Thus, WU-04 is a promising drug candidate for coronavirus treatment. One-Sentence SummaryA oral non-covalent inhibitor of 3C-like protease effectively inhibits SARS-CoV-2 replication.

microbiology↗

bioRxiv2022

Editing the CsDMR6 Gene in Citrus Results in Resistance to the Bacterial Disease Citrus Canker

Citrus is one of the most important fruit crops in the world. Citrus production worldwide faces challenges from devastating bacterial diseases, including citrus canker caused by Xanthomonas citri ssp. citri (Xcc). Improving citrus resistance to citrus canker and other major bacterial diseases has been a top priority in citrus biotechnology. Disabling disease susceptibility genes has emerged as a novel, promising approach to engineering disease resistance. The bottleneck for applying such an approach has been the identification of proper disease susceptibility-related genes in citrus. Here we show the first successful case of editing the CsDMR6 gene in citrus and obtaining strong resistance to citrus canker in six mutants in two citrus cultivars, Duncan grapefruit and Carrizo citrange. Multiple types of deletions and insertions were induced in CsDMR6, resulting in frameshift of its coding region and presumably loss of gene function. The mutation frequency in most of the mutants reached 71.8% to 98.9%. The mutants showed 71.2% to 99.8% reduction in citrus canker lesion and greater than 99.7% or 2.45 to 4.95 Log10 unit reduction in Xcc bacterial cell population. Mutants also accumulated more salicylic acid and expressed much higher levels of the NPR1 gene than the wildtype with or without Xcc inoculation, which suggests potential resistance to other diseases in these mutants. The guide RNAs for targeting CsDMR6 were selected from highly conserved regions and have 100% nucleotide identity with DMR6 homologs in important citrus species; these guide RNAs are expected to work in other important citrus species and cultivars.

plant biology↗