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Geographical background and dispersal ability may strongly influence assemblage dissimilarity; however, these aspects have generally been overlooked in previous large-scale beta diversity studies. Here, we examined whether the patterns and drivers of taxonomic beta diversity (TBD) and phylogenetic beta diversity (PBD) of breeding birds in China vary across (1) regions on both sides of the Hu Line, which demarcates China’s topographical, climatic, economic, and social patterns, and (2) species with different dispersal ability. TBD and PBD were calculated and partitioned into turnover and nestedness components using a moving window approach. Variables representing climate, habitat heterogeneity, and habitat quality were employed to evaluate the effects of environmental filtering. Spatial distance was considered to assess the impact of dispersal limitation. Variance partitioning analysis was applied to assess the relative roles of these variables. In general, the values of TBD and PBD were high in mountainous areas and were largely determined by environmental filtering. However, different dominant environmental filters on either side of the Hu Line led to divergent beta diversity patterns. Specifically, climate-driven species turnover and habitat heterogeneity-related species nestedness dominated the regions east and west of the line, respectively. Additionally, bird species with stronger dispersal ability were more susceptible to environmental filtering, resulting in more homogeneous assemblages. Our results indicated that regions with distinctive geographical backgrounds may present different ecological factors that lead to divergent assemblage dissimilarity patterns, and dispersal ability determines the response of assemblages to these ecological factors. Identifying a single universal explanation for the observed pattern without considering these aspects may lead to simplistic or incomplete conclusions. Consequently, a comprehensive understanding of large-scale beta diversity patterns and effective planning of conservation strategies necessitate the consideration of both geographical background and species dispersal ability.
Neural tube defects (NTDs) are severe congenital neurodevelopmental disorders arising from incomplete neural tube closure. Although folate supplementation has been shown to mitigate the incidence of NTDs, some cases, often attributable to genetic factors, remain unpreventable. The SHROOM3 gene has been implicated in NTD cases that are unresponsive to folate supplementation; at present, however, the underlying mechanism remains unclear. Neural tube morphogenesis is a complex process involving the folding of the planar epithelium of the neural plate. To determine the role of SHROOM3 in early developmental morphogenesis, we established a neuroepithelial organoid culture system derived from cynomolgus monkeys to closely mimic the in vivo neural plate phase. Loss of SHROOM3 resulted in shorter neuroepithelial cells and smaller nuclei. These morphological changes were attributed to the insufficient recruitment of cytoskeletal proteins, namely fibrous actin (F-actin), myosin II, and phospho-myosin light chain (PMLC), to the apical side of the neuroepithelial cells. Notably, these defects were not rescued by folate supplementation. RNA sequencing revealed that differentially expressed genes were enriched in biological processes associated with cellular and organ morphogenesis. In summary, we established an authentic in vitro system to study NTDs and identified a novel mechanism for NTDs that are unresponsive to folate supplementation.
PTEN-induced putative kinase 1 (PINK1), a mitochondrial kinase that phosphorylates Parkin and other proteins, plays a crucial role in mitophagy and protection against neurodegeneration. Mutations in PINK1 and Parkin can lead to loss of function and early onset Parkinson’s disease. However, there is a lack of strong in vivo evidence in rodent models to support the theory that loss of PINK1 affects mitophagy and induces neurodegeneration. Additionally, PINK1 knockout pigs (Sus scrofa) do not appear to exhibit neurodegeneration. In our recent work involving non-human primates, we found that PINK1 is selectively expressed in primate brains, while absent in rodent brains. To extend this to other species, we used multiple antibodies to examine the expression of PINK1 in pig tissues. In contrast to tissues from cynomolgus monkeys (Macaca fascicularis), our data did not convincingly demonstrate detectable PINK1 expression in pig tissues. Knockdown of PINK1 in cultured pig cells did not result in altered Parkin and BAD phosphorylation, as observed in cultured monkey cells. A comparison of monkey and pig striatum revealed more PINK1-phosphorylated substrates in the monkey brain. Consistently, PINK1 knockout in pigs did not lead to obvious changes in the phosphorylation of Parkin and BAD. These findings provide new evidence that PINK1 expression is specific to primates, underscoring the importance of non-human primates in investigating PINK1 function and pathology related to PINK1 deficiency.
, Available online , doi: 10.24272/j.issn.2095-8137.2023.364
The onset of the Indian monsoons has been one of the most important climate events in Asia to occur during the Cenozoic and may drive faunal and floral exchange between continents. Based on extensive phylogenetic analysis of Scytodes spitting spiders, we examine whether a Cenozoic biogeographic link between Southeast/South Asia and East/South Africa was in place or if the Indian monsoons played a role in the Asian-African faunal exchange of the genus. Additionally, we examine the global biogeography of the Scytodes thoracica group via 23 years of sampling at the species level (61 species of the thoracica group and 84 species of Scytodes) using DNA data from six loci. Our results indicate that the thoracica group initially dispersed from Southeast Asia to East Africa between 46.5 and 33.0 million years ago, and dispersal events increased between Southeast/South Asia and East/South Africa from the early to late Miocene. The timing of these events indicates that Asian-African faunal exchange of the thoracica group was driven by the Indian monsoons, and shift of the Indian monsoon from a North-South direction to an East-West direction since the middle Eocene is revealed.
A growing number of studies have identified that repeated exposure to Sevoflurane during development leads to long-term social abnormalities and cognitive impairment. Davunetide is an activity fragment of activity dependent neuroprotective protein (ADNP), which was coupled to social and cognitive protection. Whether Davunetide could attenuate the social deficits after sevoflurane exposure and the underlying developmental mechanisms are poorly understood. In this study, ribosome and proteome profiles were conducted to investigate the molecular basis in neonatal mice with sevoflurane-induced social deficits. We investigated the neuropathological basis through techniques such as Golgi staining, morphological analysis, Western blot, electrophysiology technology, and behavior analysis. Results showed that ADNP was significantly downregulated after Sevoflurane exposure during development. After adulthood, neurons in ACC of sevoflurane exhibited decrease in number of dendrites, total dendrite length and spine density. The expression of Homer, PSD95, synaptophsin and vglut2 were significantly reduced in sevoflurane group. Patch-clamp recording showed that frequency and amplitude of miniature excitatory postsynaptic currents (mEPSCs) were reduced. Importantly, davunetide significantly alleviated Sevoflurane induced synaptic defect, social behavior and cognitive impairment. Mechanistic analysis revealed that loss of ADNP caused Ca2+ activity dysregulation via Wnt/β-catenin signaling, leading to decreased the expression of synaptic proteins. Wnt suppression was all restored in davunetide-treated group. Taken together, we identified ADNP as a promising therapeutic target for the prevention and treatment of neurodevelopmental toxicity caused by general anesthetics. Our data provide an insight into the social abnormalities and cognitive damage induced by sevoflurane exposure in neonatal mice and its underlying regulatory mechanism.
Dormancy is a fascinating survival technique that enables organisms to adapt to their environment and can be classified as either hibernation or aestivation, depending on the season. Organisms that are in a state of dormancy can display remarkable stress resistance, enabling them to survive in extreme environments. However, the question of how organisms adapt to their environment in their respective dormant states and how the two types of dormancy relate to each other and differ from each other still deserves further exploration. We selected Perccottus glenii and Protopterus annectens as the primary subjects to study hibernation and aestivation respectively. We analyzed them histologically and collected multiple organ transcriptome data from both species. The functional enrichment results reveal that dormancy entails a comprehensive alteration coordinated by multiple organs. Additionally, the differences between the two dormant species (regarding coping mechanisms to deal with extreme temperatures) were considerable. We also discovered noteworthy similarities in the expression patterns of genes linked to energy metabolism, neural activity, and biosynthesis during both dormant periods, indicating a correlation between hibernation and aestivation. To comprehend the connection between the two dormant species, we scrutinised the gene homology of various other distantly related species in dormant states from public databases. We discovered that almost half of the directly homologous genes showing significant differential expression during dormancy displayed a consistent expression trend. These genes, which exhibit similar expression patterns, mainly regulate metabolism during dormancy, and maintaining the biologically dormant state requires cell number and neural activity. This suggests that the similarity between hibernation and aestivation is due to convergent evolution. In conclusion, our study enhances the comprehension of the dormancy phenomenon and offers new insights into the molecular mechanisms behind vertebrate dormancy.
The leopard coral grouper (<i>Plectropomus leopardus</i>) is a species of significant economic importance. While artificial cultivation of <i>P. leopardus</i> has thrived in recent decades, the advancement of selective breeding has been impeded by the absence of comprehensive population genome data. In this study, we identified over 8.73 million single nucleotide polymorphisms (SNPs) through whole-genome resequencing of 326 individuals spanning six distinct groups. Furthermore, we categorized 226 individuals with high-coverage sequencing (depth ≥ 14×) into eight clusters based on their genetic profiles and phylogenetic relationships. Notably, four of these clusters exhibited pronounced genetic differentiation compared with the other populations. To identify potentially advantageous loci for <i>P. leopardus</i>, we examined genomic regions exhibiting selective sweeps by analyzing the nucleotide diversity (<i>θπ</i>) and fixation index (<i>F<sub>st</sub></i>) in these four clusters. Moreover, by leveraging these high-coverage re-sequencing data, we successfully constructed the first haplotype reference panel specific to <i>P. leopardus</i>. This achievement holds promise for enabling high-quality, cost-effective imputation methods. Additionally, we used low-coverage sequencing data in conjunction with imputation for a genome-wide association study to identify candidate SNP loci and genes associated with growth traits. A significant concentration of these genes was observed on chromosome 17, which is primarily involved in skeletal muscle and embryonic development and cell proliferation. Notably, our investigation of growth-related SNPs across the eight clusters revealed that cluster 5 harbored the most promising candidate SNPs for genetic selective breeding efforts. These findings provide a robust toolkit and valuable insights into the management of germplasm resources and genome-driven breeding initiatives targeting <i>P. leopardus</i>.
, Available online , doi: 10.24272/j.issn.2095-8137.2023.236
In eukaryotes, N6-methyladenosine (m6A) is the most prevalent internal modification of messenger RNA (mRNA), and it can be reversibly regulated by specific m6A writers and erasers. FTO catalyzes RNA demethylation and plays a critical role in physiological and pathological processes. Our study found dynamic changes in both m6A and FTO during primordial follicle assembly, with opposite trends observed for m6A levels and nucleus-localized FTO expression. Fto siRNA altered the expression of genes related to cell proliferation, hormone regulation, and cell chemotaxis, and also affected RNA alternative splicing. Overexpression of full-length Fto resulted in changes to m6A levels, the ratio of alternative splicing forms of Cdk5, cell proliferation, cell cycle, and the proportion of primordial follicle. However, overexpression of Fto lacking a nuclear localization signal (NLS) had no significant effects on m6A levels or primordial follicle assembly. Our findings suggest that FTO expressed in the nucleus but not in the cytoplasm, regulates RNA m6A demethylation and is involved in cell proliferation, cell cycle and primordial follicle assembly. These results highlight the potential of m6A and its eraser FTO as biomarkers and therapeutic targets.
, Available online , doi: 10.24272/j.issn.2095-8137.2023.222
Animal body size variation is of particular interest in evolutionary biology, but the genetic basis remains largely unknown. Previous studies have shown the presence of two parallel evolutionary genetic clusters within the fish genus Epinephelus, which showed evident divergence in body size, providing an excellent opportunity to investigate the genetic basis of body size variation in vertebrates. Herein, we performed phylo-transcriptomic methods and reconstructed the phylogeny of 13 epinephelids originated from the South China Sea. Two genetic clades with estimated divergence time of ~ 15.4 MYA (million years ago), were identified to correlate with large and small body size, respectively. A total of 180 rapidly evolving genes (REGs) and two positively selected genes (PSGs) were identified between the two groups. Functional enrichment analyses on these candidate genes revealed distinctly different enrichment categories between the two groups. These pathways or genes may play important roles in body size variation in groupers through complex regulatory networks. Based on the results of this study, we speculate that the ancestors of the two divergent groups of groupers may have adapted to different environments through habitat selection, leading to genetic variations in metabolic patterns, organ development, and lifespan, which finally resulted in the divergence of body size between two locally adapted populations. These findings provide important insights into the genetic mechanisms underlying body size variation in groupers and species differentiation.
, Available online , doi: 10.24272/j.issn.2095-8137.2023.205
The placenta plays a crucial role in the successful reproduction of mammals. Ruminant animals possess a semi-invasive placenta characterized by a highly vascularized structure and formed by maternal endometrial caruncles and fetal placental cotyledons. This specialized placenta is essential for fetal development until full term. The cow placenta consists of at least two trophoblast cell populations, including uninucleate (UNC) and binucleate (BNC) cells. However, the current inability to describe the transcriptomic dynamics of the placental natural environment has resulted in a poor understanding of the molecular and cellular interactions between trophoblast cells and niches, as well as of the molecular mechanisms controlling trophoblast differentiation and functionalization. To fill in this knowledge gap, we employed Stereo-seq, a spatial transcriptomics technique, to generate a map capturing the spatial gene expression patterns at near single-cell resolution in the cow placenta on 90 and 130 days of gestation, and attained high-resolution, spatially resolved gene expression profiles of cow placenta. Based on clustering and cell marker gene expression, key transcription factors, including YBX1 and NPAS2, were revealed to regulate the heterogeneity of trophoblast cell subpopulations. Cell communication and trajectory analysis provided a framework for understanding cell-cell interactions and the differentiation of trophoblasts into BNCs in the microenvironment of the cow placenta. Differential analysis of cell trajectories identified a set of genes involved in regulation of trophoblast differentiation. Additionally, we identified spatial modules and co-variant genes that play critical roles in shaping specific tissue structures. Together, this foundational information contributes to the discovery of important biological pathways underlying the development and function of the cow placenta.
As social animals, Indo-Pacific humpback dolphins have community differentiation, but the external-internal influencing factors and spatiotemporal dynamics are not well known. Thus, we monitored the social structure variation in an endangered Indo-Pacific humpback dolphin population in Xiamen Bay, China, during two periods, namely, 2007–2010 and 2017–2019, and analyzed the influence of habitat use and individual composition. In both periods, the population showed highly similar social differentiation, and the individuals were divided into two main clusters and a small cluster. The two main clusters occupied the east and west waters but the core distribution area of the east cluster moved further eastward during 2007–2010 and 2017–2019, and the distribution shift did not change the temporal stability of the social structure or inter-association of the east cluster. The 16 identical individuals in the two periods (accounting for 51.6% and 43.2%, respectively) seemed to constitute the basic framework of the social structure and could be the main reason for the stable social structure over the past decade. However, these individuals likely played a more critical role in maintaining the social network structure in 2007–2010 than that in 2017–2019. These results suggested that the internal factors of the dominant individuals’ composition contributed more to building the social network than the external factor of habitat use change. Based on the findings, different protective measures have been proposed for two main clusters respectively.
Molecular phylogenetic relationship based on mitochondrial genomes within novel deep-sea corals (Octocorallia: Alcyonacea) insights into the slow evolution and adaptation of the extreme deep-sea environment
, Available online , doi: 10.24272/j.issn.2095-8137.2023.039
A total of 10 Alcyonacea corals were collected at depths ranging from 900 m to 1640 m by the manned submersible Shenhai Yongshi during two cruises in the South China Sea (SCS). Phylogenetic distance and average nucleotide identity (ANI) analyses of mitochondrial genomes combined with morphology examination and sclerite scanning showed that the collected samples could be assigned to four suborders - Calcaxonia, Holaxonia, Scleraxonia, and Stolonifera - which might represent 10 novel deep-sea species. The analyses of the dissimilarity of GC skew, phylogenetic distance, and ANI indicated that the evolution of Octocorallia mitochondrial sequences was slow. The nonsynonymous (Ka) and synonymous (Ks) substitution (Ka/Ks) ratios indicated that 14 protein-coding genes (PCGs) were undergoing purifying selection and that the selection pressures might be from specific deep-sea environments. The correlation analysis of median values of Ka/Ks ratio of five gene families and environmental factors showed that the genes encoding cytochrome b (cob) and DNA mismatch repair protein (mutS) might be driven by environmental factors to format deep-sea species. This study highlighted the slow evolution and adaptative mechanism of deep-sea corals in the deep ocean.
Omega-3 polyunsaturated fatty acids (n-3 PUFAs), particularly docosahexaenoic acid (22:6n-3, DHA), have been shown to play a crucial role in reproduction and reproductive health of vertebrates, including humans. However, the underlying mechanism of this phenomenon remains largely unknown. In this study, we used two zebrafish genetic models, the elovl2-/- mutant as an endogenous DHA-deficient model and the fat1 (an omega-3 desaturase encoding gene) transgenic zebrafish as an endogenous DHA-rich model, to investigate the effects of DHA on oocyte maturation and oocyte quality. Our results show that the elovl2-/- mutant zebrafish had much lower fecundity and poorer oocyte quality than the wildtype controls, while the fat1 zebrafish had higher fecundity and better oocyte quality than wildtype controls. DHA deficiency in elovl2-/- embryos led to defects of egg activation, poor microtubule stability and reduced pregnenolone levels. Further study reveals that DHA promotes pregnenolone synthesis by promoting the transcription of cyp11a1, which encodes the cholesterol side-chain cleavage enzyme, thereby stabilizing microtubule assembly during oogenesis. In turn, the hypothalamic–pituitary–gonadal (HPG) axis is enhanced by DHA. In conclusion, using two unique genetic models, our study demonstrates the endogenously synthesized DHA promotes oocyte maturation and oocyte quality by promoting pregnenolone production via transcriptional regulation of cyp11a1.
Glycogen is the most effective energy reserve for metabolism in aquatic shellfish, and also contributes to the flavor and quality of oyster. Jinjiang oyster Crassostrea ariakensis is an economically and ecologically important species in China. In this study, RNA sequencing (RNA-seq) and assay for transposase-accessible chromatin using sequencing (ATAC-seq) were performed respectively to explore the gene expression and dynamic changes of chromatin accessibility among the oysters with different glycogen contents. A total of 9483 differentially expressed genes (DEGs) and 7215 significantly differential chromatin accessibility genes (DCAGs) were obtained, with the intersection of DEGs and DCAGs reaching 2600. Many of those genes were enriched in the pathways related to glycogen metabolism such as "Glycogen metabolic process" and "Starch and sucrose metabolism". In addition, a total of 526 SNP loci associated with glycogen content obtained by the genome wide association study (GWAS) corresponded to 241 genes, 63 of which were also DEGs and DCAGs as revealed above. This study will enrich basic research data and provide insights into the molecular mechanisms underlying the regulation of glycogen metabolism in oyster.
, Available online , doi: 10.24272/j.issn.2095-8137.2023.012
Monitoring the prevalence of the antimicrobial resistance gene (ARG) is critical to address the global crisis of antibiotic resistant bacterial infections. However, the characterization of ARG and microbiome structure, as well as the indicators for routine ARG monitoring in pig farms, are still lacking under the variation in antimicrobial exposure between countries/regions. Hence, metagenomics and Random Forest machine learning algorithm were used to decipher the ARG profiles, microbiome, and ARG contamination indicators in pig manure under different antimicrobial pressures between China and Europe. The results showed that Chinese pigs exposed to high level antimicrobials had higher total and plasmid-mediated ARG abundances than European pigs (P < 0.05). ANT(6)-Ib, APH(3')-IIIa, and tet(40) were the shared core ARGs between Chinese and European pigs. The core ARG identified in pigs existed a linkage between corresponding country/regions pigs and humans. Moreover, Lactobacillus and Prevotella were the dominant phyla in the core microbiome of Chinese and European pigs, respectively. Forty ARG markers and 43 biomarkers were found to differentiate Chinese and European pig manure samples with 100% and 98.7% accuracy, respectively. We identified indicators to assess the ARG contamination in Chinese and European pigs with high accuracy (r = 0.72 ~ 0.88). Escherichia flexneri in Chinese and European pigs carried numerous ARGs, ranging from 21 to 37. This study emphasized the importance of global collaboration in reducing antimicrobial resistance (AMR) risk and provided indicators for evaluating the risk of ARG contamination in pig farms.
Pig H3K4me3, H3K27ac and gene expression profiles reveal reproductive tissue-specific activity of transposable elements
, Available online , doi: 10.24272/j.issn.2095-8137.2023.060
Regulatory sequences and transposable elements (TEs) account for a large proportion of the genome sequences of species, while their roles in gene transcription, especially tissue-specific expression, remain largely unknown. Pigs are an excellent animal models for studying the biology of the genome sequences owing to their great diversity of wild and domesticated populations. Here, we integrated H3K27ac ChIP-seq, H3K4me3 ChIP-seq and RNA-seq data from 10 tissues of the same 7 fetuses and their consanguineously related 8 adult pigs to annotate the regulatory elements and TEs for their links with histone modifications and mRNA expression across varying tissues and development stages. The association analyses of mRNA expression with H3K27ac and H3K4me3 peak activity revealed that H3K27ac peaks showed stronger associations with gene expression than H3K4me3. We revealed that 1.45% of the TEs overlapped with H3K27ac or H3K4me3 peaks, of which the majority displayed tissue-specific activity. We identified a TE subfamily (LTR4C_SS) with binding motifs for SIX1 and SIX4 that showed specific enrichment in adult and fetal ovary H3K27ac peaks. We also revealed widespread expression of TEs as part of exons or promoters of genes from the RNA-seq data, including 4688 TE-containing transcripts that displayed development stage and tissue-specific expression. Notably, 1967 TE-containing transcripts were enriched in the testes. We highlighted that an LTR acting as a testis-specific alternative promoter in SRPK2 (a cell cycle-related protein kinase) in our pig data, MLT1F1, was also conserved in humans and mice, suggesting an ancient embedding of the TEs in testis-specific expressed genes or parallel evolution. Collectively, our work demonstrates that TEs are deeply embedded in the genome and exhibit important tissue-specific biological functions, particularly in the reproductive organs.
, Available online , doi: 10.24272/j.issn.2095-8137.2023.286
Drug addiction is a complex and chronic disease that affects the brain and behavior, leading to an inability to control the use of substances. It is characterized by compulsive drug-seeking, drug use, and a strong desire to use the substance, as well as the development of tolerance and withdrawal. Microglia, a type of glial cell in the central nervous system (CNS), play a crucial role in maintaining the health and function of the CNS. Accumulating evidence indicates that microglia play an important role in the progression of drug addiction. From a neuroimmunopharmacological perspective, herein we discuss the role and underlying mechanisms of microglia and potential therapeutic strategies by targeting microglia for drug addiction, and the limitations of the research on microglia in drug addiction.
, Available online , doi: 10.24272/j.issn.2095-8137.2023.231
Hereditary hearing loss (HHL) is a genetic disorder that impairs auditory function, and it can seriously affect the quality of human life and lead to major economic losses for society. To investigate the underlying causes of HHL and evaluate therapeutic outcomes, appropriate animal models are necessary. Recently, pigs have been widely used as valuable large model animals in biomedical research. In this review, we highlight the advantages of pig models in terms of ear anatomy, inner ear morphology and electrophysiological characteristics, as well as the recent progress in establishing distinct genetically modified porcine models of hearing loss. Additionally, we present the prospects, obstacles and recommendations in using pig models for HHL research. This review should provide insights and perspectives for the future research of porcine models for HHL.
Animal models are extensively used in all prospects of biomedical research, making significant contributions to disease investigations, drug development, and gene function studies. As is typical with experimental animals, genome modification in rabbits has progressed slowly over the years. However, recent advancements, particularly in CRIPR/Cas9-related technologies, have catalyzed the successful development of numerous genome-edited rabbit models to effectively mimic diverse diseases, including cardiovascular disorders, immunodeficiencies, aging-related ailments, neurological diseases, ophthalmic pathologies, and others. These genome-edited rabbit models hold great promise in advancing biomedical research due to their closer physiological and biochemical resemblance to humans compared to mice. In this review, we summarize the novel gene-editing approaches available for genome-edited rabbits and present the applications and prospects of genome-edited rabbit models in biomedicine with the goal of underscoring their particular impact and future potential in translational medicine.
, Available online , doi: 10.24272/j.issn.2095-8137.2023.199
Huntington's disease (HD) is an inherited neurodegenerative disorder for which there is currently no effective treatment available. Thus, it is imperative to establish appropriate disease models to delve deeply into the comprehensive progression of the disease. The genetic cause of HD is the abnormal expansion of CAG repeats in the huntingtin (HTT) gene, which leads to the expansion of a polyglutamine repeat in the HTT protein. Mutant HTT carrying the expanded polyglutamine repeat becomes misfolded and aggregates in the brain, causing selective loss of neurons in specific brain regions. Animal models play an important role in elucidating the pathogenesis of neurodegenerative disorders, including HD, and identifying potential therapeutic targets. Given the considerable species differences between rodents and large animals, large animal models of HD have been established to investigate the HD pathogenesis. These should facilitate the discovery of novel therapeutic targets, effective drug delivery, and improve treatment outcomes. We have previously explored the advantages of utilizing large animal models, particularly pigs, in other review articles. Since then, significant progress has been made in developing more sophisticated animal models that faithfully replicate the typical pathology of HD. The current review aims to provide a more comprehensive overview of large animal models of HD, incorporating recent findings regarding the establishment of HD knock-in (KI) pigs and their genetic therapy. In this review, we also explore the utilization of large animal models in Huntington's disease (HD) research, specifically focusing on sheep, non-human primates, and pigs. Our objective is to provide valuable insights into the application of these large animal models for investigating and treating neurodegenerative disorders.