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Hsieh, C.

Publications and source records attributed to Hsieh, C..

16 recordsLinked to original sources

PKMζ-KIBRA interactions, molecular turnover, and memory

How can the molecules that strengthen synaptic connections maintain memory in the face of molecular turnover? Our previous work showed that persistent interaction between the postsynaptic scaffolding protein, KIBRA, and the autonomously active PKC isoform, PKM{zeta}, is crucial for maintaining synaptic long-term potentiation (LTP) and memory for at least a month. This duration is longer than the lifespans of individual KIBRA and PKM{zeta} molecules. Biophysical modeling of the interaction suggests oligomers of KIBRA-PKM{zeta} dimers, but not individual dimers or monomers, can overcome molecular turnover by continually incorporating newly synthesized KIBRA and PKM{zeta}, replacing those that have degraded. Here we used AlphaFold 3 to predict the structures of KIBRA-PKM{zeta} heterodimers and heterohexamers and to examine the sites of action of two structurally distinct inhibitors of KIBRA-PKM{zeta} interaction that disrupt established late-LTP and long-term memory. The structures predict that the peptide K-ZAP blocks formation of heterodimers, whereas the small molecule {zeta}-stat prevents PKM{zeta} of one heterodimer from binding a second KIBRA and PKM{zeta}, essential for forming larger oligomeric structures. We show that {zeta}-stat, like K-ZAP, disrupts 1-month-old spatial memory. Thus, continual formation of KIBRA-PKM{zeta} oligomers can be a core molecular mechanism driving the persistence of long-term memory in the face of molecular turnover.

neuroscience↗

Abundance redistribution increases predator-prey interaction potentials among North American birds

Species are dramatically shifting their geographical ranges, yet the consequences of these spatial redistributions on species interactions have not been examined at broad spatial and temporal scales. Here, we investigate how the redistribution of species abundance over the past 50 years affects the potential for predator-prey interactions among North American birds. We first predict potential predator-prey interactions based on existing data and functional traits, then construct a novel, abundance-weighted metric of interaction potential for each pair of predator and prey in local communities. Across North America, the potential for predator-prey interactions has been increasing over time. We further show that this widespread increasing trend in interaction potentials is mostly driven by higher abundances of predator species. The spatial redistributions of predators and prey lead to turnover of interactions within local communities, and the turnover is less associated with changes in climate but more associated with human-induced land use changes. Our novel approach and results highlight the importance of considering traits, abundance, and species interactions--including known and potential interactions--when studying the ecological consequences of species range shifts.

ecology↗

PKMζ-PKC{iota}/{lambda} double-knockout reveals the atypical PKCs are crucial for hippocampal late-LTP and spatial long-term memory

PKM{zeta}, a persistently active atypical PKC (aPKC) isoform, is thought to maintain late-phase long-term potentiation (late-LTP) and long-term memory. However, PKM{zeta}-knockout mice still exhibit hippocampal LTP and spatial memory while lacking neocortical LTP, questioning whether this kinase is fundamental to enduring synaptic potentiation and memory. Tsokas et al. (2016) showed the other aPKC, PKC{iota}/{lambda}, likely compensates for PKM{zeta} during maintenance in the hippocampus of PKM{zeta}-null mice. In wild-type mice, PKC{iota}/{lambda} drives early-LTP and short-term memory, while PKM{zeta} compensates for PKC{iota}/{lambda} knockout by supporting both early- and late-phase processes. Here we show PKC{iota}/{lambda} persistently increases during maintenance in two mouse models: PKM{zeta}-conditional knockout (cKO) mice, and double-knockout mice carrying both conditional deletion of PKC{iota}/{lambda} and constitutive loss of PKM{zeta}. In the double-knockout mice, PKC{iota}/{lambda} was measured while the kinase was still present, prior to its inducible ablation, to characterize its compensatory upregulation in late-LTP before removal. To test whether this compensation was functional, we ablated PKC{iota}/{lambda} in the hippocampus of the double-knockout mice. The double-knockout eliminated late-LTP, whereas individual knockout of either aPKC alone showed normal-appearing LTP. Double-knockout also abolished spatial long-term memory without affecting short-term memory. Thus, when PKM{zeta} is absent, PKC{iota}/{lambda} persists to maintain hippocampal late-LTP and long-term memory.

neuroscience↗

An updated sulfate transporter phylogeny uncovers a perennial-specific subgroup associated with lignification

Sulfate-proton co-transporters (SULTRs) mediate sulfate uptake, transport, storage, and assimilation in plants. The SULTR family has historically been classified into four groups (SULTR1-SULTR4), with well-characterized roles for SULTR groups 1, 2, and 4. However, the functions of the large and diverse SULTR3 group remain poorly understood. Here, we present an updated phylogenetic analysis of SULTRs across angiosperms, including multiple early-divergent lineages. Our results suggest that the enigmatic SULTR3 group comprises four distinct subfamilies that predate the emergence of angiosperms, providing a basis for reclassifying the SULTR family into seven subfamilies. This expanded classification is supported by subfamily-specific gene structures and amino acid substitutions in the substrate-binding pocket. Structural modeling identified three serine residues uniquely lining the substrate-binding pocket of SULTR3.4, enabling three hydrogen bonds with the phosphate ion. The data support the proposed neofunctionalization of this subfamily for phosphate allocation within vascular tissues. Transcriptome analysis of Populus tremula x alba revealed divergent tissue expression preferences among SULTR subfamilies and between genome duplicates. We observed partitioned expression in vascular tissues among the four SULTR3 subfamilies, with PtaSULTR3.4a and PtaSULTR3.2a preferentially expressed in primary and secondary xylem, respectively. Gene coexpression analysis revealed coordinated expression of PtaSULTR3.4a with genes involved in phosphate starvation responses and nutrient transport, consistent with a potential role in phosphate homeostasis. In contrast, PtaSULTR3.2a was strongly coexpressed with lignification and one-carbon metabolism genes and their upstream transcription regulators. PtaSULTR3.2a belongs to a eudicot-specific branch of the SULTR3.1 subfamily found only in perennial species, suggesting a specialized role in lignifying tissues. Together, our findings provide a refined phylogenetic framework for the SULTR family and suggest that the expanded SULTR3 subfamilies have undergone neofunctionalization during the evolution of vascular and perennial plants.

plant biology↗

Populus VariantDB v3.2 facilitates CRISPR and Functional Genomics Research

The success of CRISPR genome editing studies depends critically on the precision of guide RNA (gRNA) design. Sequence polymorphisms in outcrossing tree species pose design hazards that can render CRISPR genome editing ineffective. Despite recent advances in tree genome sequencing with haplotype resolution, sequence polymorphism information remains largely inaccessible to various functional genomics research efforts. The Populus VariantDB v3.2 addresses these challenges by providing a user-friendly search engine to query sequence polymorphisms of heterozygous genomes. The database accepts short sequences, such as gRNAs and primers, as input for searching against multiple poplar genomes, including hybrids, with customizable parameters. We provide examples to showcase the utilities of VariantDB in improving the precision of gRNA or primer design. The platform-agnostic nature of the probe search design makes Populus VariantDB v3.2 a versatile tool for the rapidly evolving CRISPR field and other sequence-sensitive functional genomics applications. The database schema is expandable and can accommodate additional tree genomes to broaden its user base.

plant biology↗

Uncovering synaptic and cellular nanoarchitecture of brain tissue via seamless in situ trimming and milling for cryo-electron tomography

Cell-cell communication underlies all emergent properties of the brain, including cognition, learning and memory. The physical basis for these communications is the synapse, a multi-component structure requiring coordinated interactions between diverse cell types. However, many aspects of three-dimensional (3D) synaptic organization remain poorly understood. Here, we developed an approach, seamless in situ trimming and milling (SISTM), to reliably fabricate sufficiently thin lamellae for mapping of the 3D nanoarchitecture of synapses in mouse, monkey and human brain tissue under near-native conditions via cryo-electron tomography (cryo-ET). We validated SISTM in a mouse model of Huntingtons disease, demonstrating distinct 3D alterations to synaptic vesicles and mitochondria. By successfully applying SISTM to macaque brain, we described the 3D architecture of a tripartite synapse within the cortex. Subtomogram averaging (STA) enabled spatial mapping of astrocyte-neuron contacts within the tripartite synapse, revealing neurexin-neuroligin complexes as potential constituents that tether the two cell types. Finally, we showed that the defining features of synaptic nanoarchitecture were conserved across species and evident in human brain tissue obtained postmortem. Combining SISTM with cryo-ET and STA is a starting point for a new understanding of brain organization, disease-induced structural alterations and the development of rational, structure-guided therapeutics.

cell biology↗

Increased protein kinase Mζ expression by Minocycline and N-acetylcysteine restoreslate-phase long-term potentiation and spatial learning after closed head injury in mice

Cognitive deficits frequently arise after traumatic brain injury. The murine closed head injury (CHI) models these deficits since injured mice cannot acquire Barnes maze. Dosing of minocycline plus N-acetylcysteine beginning 12 hours post-CHI (MN12) restores Barnes maze acquisition by an unknown mechanism. Increased hippocampal synaptic efficacy is needed to acquire Barnes maze, synaptic long-term potentiation (LTP) models this increased synaptic efficacy in vitro. LTP has an early phase (E-LTP) lasting up to one hour that is mediated by second messengers that is followed by a late phase (L-LTP) that needs new synthesis of protein kinase M zeta (PKM{zeta}). PKM{zeta} has constitutive kinase activity because it lacks the autoinhibitory regulatory domain found in other PKCs. Due to its constitutive activity, the amount of PKM{zeta} kinase activity is determined by PKM{zeta} protein levels. We report that CHI bilaterally decreases PKM{zeta} levels in the CA3 and CA1 hippocampus. MN12 increases CA1 PKM{zeta} expression. CHI inhibits E-LTP in slices from the ipsilesional hippocampus and inhibits L-LTP in slices from both hippocamppi. MN12 treatment reestablishes both E-LTP and L-LTP in slices from the injured MN12-treated hippocampus. The restoration of L-LTP from injured MN12-treated hippocampus is mediated by PKM{zeta} because L-LTP is blocked by the specific PKM{zeta} inhibitor, {zeta}-stat. Hippocampal {zeta}-stat infusions also prevents Barnes maze acquisition in injured, MN12-treated mice. These data suggest that post-injury minocycline plus N-acetylcysteine targets PKM{zeta} to improve synaptic plasticity and cognition in mice with closed-head injury.

neuroscience↗

Amygdala stimulation transforms short-term memory into remote memory by persistent activation of atypical PKC in the anterior cingulate cortex

Although many studies have addressed the role of the amygdala in modulating long-term memory, it is not known whether weak training plus amygdala stimulation can transform a short-term memory into a remote memory. Object place recognition (OPR) memory after strong training remains hippocampus-dependent through the persistent action of PKM{zeta} for at least 6 days, but it is unknown whether weak training plus amygdala stimulation can transform short-term memory into an even longer memory, and whether such memory is stored through more persistent action of PKM{zeta} in hippocampus. We trained rats to acquire OPR and 15 min or 5 h later induced a brief pattern of electrical stimulation in basolateral amygdala (BLA). Our results reveal that a short-term memory lasting < 4 h can be converted into remote memory lasting at least 3 weeks if the BLA is activated 15 min, but not 5 h after learning. To examine how this remote memory is maintained, we injected ZIP, an inhibitor of atypical PKCs (aPKCs), PKM{zeta} and PKC{iota}/{lambda}, into either hippocampal CA1, dentate gyrus (DG), or anterior cingulate cortex (ACC). Our data reveal amygdala stimulation produces consolidation into remote memory, not by persistent aPKC activation and capture by synaptic tagging processes in the hippocampal formation, but in ACC. Our data establish a powerful modulating role of the BLA in forming remote memory and open a path in the search for neurological restoration of memory, based on enhancing synaptic plasticity in aging or neurodegenerative disorders such as Alzheimers disease.

neuroscience↗

Counter-regulation of RNA stability by UPF1 and TDP43

RNA quality control is crucial for proper regulation of gene expression. Disruption of nonsense mediated mRNA decay (NMD), the primary RNA decay pathway responsible for the degradation of transcripts containing premature termination codons (PTCs), can disrupt development and lead to multiple diseases in humans and other animals. Similarly, therapies targeting NMD may have applications in hematological, neoplastic and neurological disorders. As such, tools capable of accurately quantifying NMD status could be invaluable for investigations of disease pathogenesis and biomarker identification. Toward this end, we assemble, validate, and apply a next-generation sequencing approach (NMDq) for identifying and measuring the abundance of PTC-containing transcripts. After validating NMDq performance and confirming its utility for tracking RNA surveillance, we apply it to determine pathway activity in two neurodegenerative diseases, amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) characterized by RNA misprocessing and abnormal RNA stability. Despite the genetic and pathologic evidence implicating dysfunctional RNA metabolism, and NMD in particular, in these conditions, we detected no significant differences in PTC-encoding transcripts in ALS models or disease. Contrary to expectations, overexpression of the master NMD regulator UPF1 had little effect on the clearance of transcripts with PTCs, but rather restored RNA homeostasis through differential use and decay of alternatively poly-adenylated isoforms. Together, these data suggest that canonical NMD is not a significant contributor to ALS/FTD pathogenesis, and that UPF1 promotes neuronal survival by regulating transcripts with abnormally long 3UTRs.

neuroscience↗

Host brain environmental influences on transplanted medial ganglionic eminence progenitors

Interneuron progenitor transplantation can ameliorate disease symptoms in a variety of neurological disorders. This strategy is based on transplantation of embryonic medial ganglionic eminence (MGE) progenitors. Elucidating host brain environment influences on interneuron progenitors as they integrate is critical to optimizing this strategy across different disease states. Here, we systematically evaluated age and brain region influences on survival, migration and differentiation of transplant-derived cells. We find that early postnatal MGE transplantation yields superior survival and more extensive migratory capabilities compared to juvenile or adult. MGE progenitors migrate more widely in cortex compared to hippocampus. Maturation to interneuron subtypes is regulated by age and brain region. MGE progenitors transplanted into dentate gyrus sub-region of early postnatal hippocampus can differentiate into astrocytes. Our results suggest that host brain environment critically regulates survival, spatial distribution and maturation of MGE-derived interneurons following transplantation. These findings inform and enable optimal conditions for interneuron transplant therapies.

neuroscience↗

Sucrose or starch? The influence of tonoplast sucrose transporter perturbation on carbon partitioning for growth, defense, and winter protection in coppiced poplar

Non-structural carbohydrate reserves of stems and roots underpin overall tree fitness. These reserves also contribute to productivity under short-rotation management practices such as coppicing for bioenergy. While sucrose and starch comprise the predominant carbohydrate reserves of Populus, utilization is understood primarily in terms of starch turnover. The tonoplast sucrose transport protein SUT4 modulates sucrose export from source leaves to distant sinks during photoautotrophic growth, but the possibility of its involvement in remobilizing carbohydrates from sinks during heterotrophic growth has not been explored. Here, we used PtaSUT4-knockout mutants of Populus tremula x P. alba (INRA 717-1B4) in winter (cool) and summer (warm) glasshouse coppicing experiments to strain carbon demand and test for SUT4 involvement in reserve utilization. We show that epicormic bud emergence, subsequent growth, and xylem hexose increases were delayed or reduced in sut4 mutants following lower temperature winter coppicing. Depletion of stem reserves during post-coppice regrowth was not impaired in sut4, but sut4 winter maintenance costs may have been higher in metabolic terms. Bark accrual of abundant defense metabolites, including salicinoids, chlorogenic acids, and flavonoid products, was prioritized in the summer, but attenuated in sut4 mutants. Summer sprout growth was not reduced in sut4 compared to controls. Together, our results point to shifting priorities for SUT4 modulation of seasonal trade-offs between growth and other priorities in the stem and emerging buds during reserve utilization in Populus.

plant biology↗

Cryo-FIB workflow for imaging brain tissue via in situ cryo-electron microscopy

Cryo-electron microscopy (cryo-EM) enables the study of protein complexes, cytoskeletal elements, and organelles in three dimensions without the use of chemical fixation. Most cryo-EM studies focus on vitreously frozen individual cells separated from their native tissue contexts. This reliance on imaging of single cells is primarily due to technical challenges associated with preparing fresh tissue sections at a thinness sufficient for visualization via cryo-EM. Highly heterogenous and specialized tissues, such as brain, are especially affected by this limitation as the cellular, subcellular, and synaptic milieus can significantly vary across neuroanatomical locations. To address this limitation, we established new instrumentation and a workflow that consists of: 1) high-pressure freezing of fresh brain tissue; 2) tissue trimming followed by cryo-focused ion beam milling via the H-bar approach to generate ultrathin lamellae; and 3) cryo-EM imaging. Here, we apply this workflow to visualize the fine ultrastructural details of organelles, as well as cytoskeletal and synaptic elements that comprise the cortical neuropil within fresh, unfixed mouse brain tissue. Moreover, we present initial studies that apply principles of the above workflow to the analysis of postmortem human brain tissue. Overall, our work integrates the strengths of cryo-electron microscopy and tissue-based approaches to produce a generalizable workflow capable of visualizing subcellular structures within complex tissue environments.

neuroscience↗

MdrDB: Mutation-induced drug resistance DataBase

AO_SCPLOWBSTRACTC_SCPLOWMutation-induced drug resistance - where the efficacy of drugs is diminished by structural changes in proteins - presents a significant challenge to drug development and the clinical treatment of disease. Understanding the effects of mutation on protein-ligand binding affinities is a key step in developing more effective drugs and therapies, but as a research community we are currently hindered by the lack of a comprehensive database of relevant information. To address this issue, we have developed MdrDB, a database of information related to changes in protein-ligand affinity caused by mutations in protein structure. MdrDB combines data from seven publicly available datasets with calculated biochemical features, as well as 3D structures computed with PyMOL and AlphaFold 2.0, to form the largest database of its kind. With 3D structural information provided for all samples, MdrDB was specifically created to have the size, breadth, and complexity to be useful for practical protein mutation studies and drug resistance modeling. The database brings together wild type and mutant protein-ligand complexes, binding affinity changes upon mutation ({Delta}{Delta}G), and biochemical features calculated from complexes to advance our understanding of mutation-induced drug resistance, the development of combination therapies, and the discovery of novel chemicals. In total, MdrDB contains 100,537 samples generated from 240 proteins (5,119 total PDB structures), 2,503 mutations, and 440 drugs. Of the total samples, 95,971 are based on available PDB structures, with the remaining 4,566 based on AlphaFold 2.0 predicted structures.

bioinformatics↗

Effects of sub-lethal single, simultaneous, and sequential abiotic stresses on phenotypic traits of Arabidopsis thaliana

Plant responses to abiotic stresses are complex and dynamic, and involve changes in different traits, either as the direct consequence of the stress, or as an active acclimatory response. Abiotic stresses frequently occur simultaneously or in succession, rather than in isolation. Despite this, most studies have focused on a single stress and single or few plant traits. To address this gap, our study comprehensively and categorically quantified the individual and combined effects of three major abiotic stresses associated with climate change (flooding, progressive drought and high temperature) on 12 phenotypic traits related to morphology, development, growth and fitness, at different developmental stages in four Arabidopsis thaliana accessions. Combined sub-lethal stresses were applied either simultaneously (high temperature and drought) or sequentially (flooding followed by drought). In total, we analyzed the phenotypic responses of 1782 individuals across these stresses and different developmental stages. Overall, abiotic stresses and their combinations resulted in distinct patterns of effects across the traits analyzed, with both quantitative and qualitative differences across accessions. Stress combinations had additive effects on some traits, whereas clear positive and negative interactions were observed for other traits: 9 out of 12 traits for high temperature and drought, 6 out of 12 traits for post-submergence and drought showed significant interactions. In many cases where the stresses interacted, the strength of interactions varied across accessions. Hence, our results indicated a general pattern of response in most phenotypic traits to the different stresses and stress combinations, but it also indicated a natural genetic variation in the strength of these responses. Overall, our study provides a rich characterization of trait responses of Arabidopsis plants to sub-lethal abiotic stresses at the phenotypic level and can serve as starting point for further in-depth physiological research and plant modelling efforts.

plant biology↗

Deep-BGCpred: A unified deep learning genome-mining framework for biosynthetic gene cluster prediction

Natural products produced by microorganisms constitute an important source of essential pharmaceuticals, including antimicrobial and anti-tumor drugs. These bioactive molecules are microbial secondary metabolites synthesized by co-localized genes termed Biosynthetic Gene Clusters (BGCs). The rapid increase of microbial genomics resources, due to the availability of high-throughput sequencing technologies, has spurred the development of computational methods for microbial genome mining for BGC discovery. Current machine learning methods, however, have limited successes in uncovering novel BGCs due to an excessive number of false positives in their predictions. To this end, we propose Deep-BGCpred, a framework that effectively addresses the aforementioned issue by improving a deep learning model termed DeepBGC. The new model embeds multi-source protein family domains and employs a stacked Bidirectional Long Short-Term Memory model to boost accuracy for BGC identifications. In particular, it integrates two customized strategies, sliding window strategy and dual-model serial screening, to improve the models performance stability and reduce the number of false positive in BGC predictions. We compare the proposed model against other well-established methods on common benchmarks and achieve new state-of-the-art results with convincing evidences. We expect that researchers working on genome mining for natural products may be greatly benefited from our newly proposed method, Deep-BGCpred.

bioinformatics↗

PKMζ traces hippocampal LTP maintenance and spatial long-term memory

PKM{zeta} is an autonomously active PKC isoform crucial for the maintenance of synaptic long-term potentiation (LTP) and long-term memory. Unlike other protein kinases that are transiently stimulated by second messengers, PKM{zeta} is persistently activated through sustained increases in kinase protein expression. Therefore, visualizing increases in PKM{zeta} expression during long-term memory storage might reveal the sites of its persistent action and thus the location of memory-associated LTP maintenance in the brain. Using quantitative immunohistochemistry validated by the lack of staining in PKM{zeta}-null mice, we examined the amount and distribution of PKM{zeta} in subregions of the hippocampal formation of wild-type mice during LTP maintenance and spatial long-term memory storage. During LTP maintenance in hippocampal slices, PKM{zeta} increases in the pyramidal cell body and stimulated dendritic layers of CA1 for at least 2 h. During spatial memory storage, PKM{zeta} increases in CA1 pyramidal cells for at least 1 month, paralleling the persistence of the memory. The subset of CA1 pyramidal cells that are tagged by immediate early gene Arc-driven transcription of fluorescent proteins, whose expression increases during initial memory formation, also expresses the persistent increase of PKM{zeta} during memory storage. In the memory-tagged cells, the increased PKM{zeta} expression persists in dendritic compartments within stratum radiatum for 1 month, indicating the long-term storage of information in the CA3-to-CA1 pathway during remote spatial memory. We conclude that persistent increases in PKM{zeta} trace the molecular mechanism of LTP maintenance and thus the sites of information storage within brain circuitry during long-term memory.

neuroscience↗