Biology subjects

Pan, M.

Publications and source records attributed to Pan, M..

116 records · Page 7Linked to original sources

bioRxiv2020

Structure of the teneurin-latrophilin complex: Alternative splicing controls synapse specificity by a novel mechanism

The trans-synaptic interaction of the cell-adhesion molecules teneurins (Tenms) with latrophilins (Lphns) promotes excitatory synapse formation when Lphns simultaneously interact with FLRTs. Insertion of a short alternatively-spliced region within Tenms abolishes the Tenm-Lphn interaction and switches Tenm function to specify inhibitory synapses. How Tenms bind to Lphns in a manner regulated by alternative splicing remains unclear. Here, we report the high-resolution cryo-EM structure of the Tenm2-Lphn3 complex, and describe the trimeric Tenm2-Lphn3-FLRT3 complex. The structure reveals that the N-terminal lectin-like domain of Lphn3 binds to the Tenm2 barrel at a site far away from the alternatively-spliced region. Alternative-splicing regulates the Tenm2-Lphn3 interaction by hindering access to the Lphn-binding surface rather than altering it. Strikingly, mutagenesis of the Lphn-binding surface of Tenm2 abolishes the Lphn3 interaction and impairs excitatory but not inhibitory synapse formation. These results suggest that a multi-level coincident binding mechanism mediated by a cryptic adhesion complex between Tenms and Lphns regulates synapse specificity.

biochemistry↗

bioRxiv2020

The Gossypium longicalyx genome as a resource for cotton breeding and evolution

Cotton is an important crop that has made significant gains in production over the last century. Emerging pests such as the reniform nematode have threatened cotton production. The rare African diploid species Gossypium longicalyx is a wild species that has been used as an important source of reniform nematode immunity. While mapping and breeding efforts have made some strides in transferring this immunity to the cultivated polyploid species, the complexities of interploidal transfer combined with substantial linkage drag have inhibited progress in this area. Moreover, this species shares its most recent common ancestor with the cultivated A-genome diploid cottons, thereby providing insight into the evolution of long, spinnable fiber. Here we report a newly generated de novo genome assembly of G. longicalyx. This high-quality genome leveraged a combination of PacBio long-read technology, Hi-C chromatin conformation capture, and BioNano optical mapping to achieve a chromosome level assembly. The utility of the G. longicalyx genome for understanding reniform immunity and fiber evolution is discussed.

genomics↗

bioRxiv2019

Regeneration of dorsal spinal cord neurons after injury via in situ NeuroD1-mediated astrocyte-to-neuron conversion

Spinal cord injury (SCI) often leads to impaired motor and sensory functions, partially because the injury-induced neuronal loss cannot be easily replenished through endogenous mechanisms. In vivo neuronal reprogramming has emerged as a novel technology to regenerate neurons from endogenous glial cells by forced expression of neurogenic transcription factors. We have previously demonstrated successful astrocyte-to-neuron conversion in mouse brains with injury or Alzheimers disease by overexpressing a single neural transcription factor NeuroD1 via retroviruses. Here we demonstrate regeneration of dorsal spinal cord neurons from reactive astrocytes after SCI via adeno-associated virus (AAV), a more clinically relevant gene delivery system. We find that NeuroD1 converts reactive astrocytes into neurons in the dorsal horn of stab-injured spinal cord with high efficiency ([~]95%). Interestingly, NeuroD1-converted neurons in the dorsal horn mostly acquire glutamatergic neuronal subtype, expressing spinal cord-specific markers such as Tlx3 but not brain-specific markers such as Tbr1, suggesting that the astrocytic lineage and local microenvironment affect the cell fate of conversion. Electrophysiological recordings show that the NeuroD1-converted neurons can functionally mature and integrate into local spinal cord circuitry by displaying repetitive action potentials and spontaneous synaptic responses. We further show that NeuroD1-mediated neuronal conversion can occur in the contusive SCI model, allowing future studies of evaluating this reprogramming technology for functional recovery after SCI. In conclusion, this study may suggest a paradigm shift for spinal cord repair using in vivo astrocyte-to-neuron conversion technology to generate functional neurons in the grey matter.

neuroscience↗

bioRxiv2019

Molecular Mechanism of Action of Mitochondrial Therapeutic SS-31 (Elamipretide): Membrane Interactions and Effects on Surface Electrostatics

Mitochondrial dysfunction includes heritable diseases, acquired pathologies, and age-related declines in health. Szeto-Schiller (SS) peptides comprise a class of amphipathic tetrapeptides that have demonstrated efficacy in treating a wide array of mitochondrial disorders, and are believed to target mitochondrial membranes due to their enrichment in the anionic phospholipid cardiolipin (CL). However, little is known regarding how SS peptides interact with or alter the physical properties of lipid bilayers. In this study, we have analyzed the interactions of the lead compound SS-31 (Elamipretide) with model and mitochondrial membranes using biophysical and computational approaches. Our results show that this polybasic peptide partitions into the membrane interfacial region with affinity and binding density that are directly related to surface charge. SS-31 binding does not destabilize lamellar bilayers even at the highest binding concentrations; however, it does cause saturable alterations in lipid packing. Most notably, SS-31 modulates the surface electrostatic properties of model and mitochondrial membranes, which could play a significant role in the mitoprotective properties of this compound. As a proof of concept, we show that SS-31 alters ion distribution at the membrane interface with implications for maintaining mitochondrial membranes subject to divalent cation (calcium) stress. Taken together, these results support a mechanism of action in which SS peptides interact with lipid bilayers and alter the biophysical (primarily electrostatic) properties of mitochondrial membranes as their primary mechanism of action. Understanding this molecular mechanism is key to the development of future compound variants with enhanced efficacy.\n\nSignificanceSzeto-Schiller (SS) peptides are among the most promising therapeutic compounds for mitochondrial dysfunction. However, the molecular target(s) and the mechanism of action of SS peptides are poorly understood. In this study, we evaluate the interaction of the lead compound SS-31 (Elamipretide) with mitochondrial and synthetic model membranes using a host of biophysical techniques. Our results show that SS-31 membrane interaction is driven largely by the negative surface charge of mitochondrial membranes and that SS-31 alters lipid bilayer properties, most notably electrostatics at the membrane interface. This work supports a mechanism in which SS peptides act on a key physical property of mitochondrial membranes rather than with a specific protein complex, consistent with the exceptionally broad therapeutic efficacy of these compounds.

biophysics↗

bioRxiv2019

Functional divergence of two duplicated Fertilization Independent Endosperm genes in rice with respect to seed development

Fertilization Independent Endosperm (FIE) is an essential member of Polycomb Repression Complex 2 (PRC2) that plays important roles in the developmental regulation of plants. OsFIE1 and OsFIE2 are two FIE homologs in the rice genome. Here, we showed that OsFIE1 probably duplicated from OsFIE2 after the origin of the tribe Oryzeae, but has a specific expression pattern and methylation landscape. During evolution, OsFIE1 underwent a less intensive purifying selection than did OsFIE2. The mutant osfie1 produced smaller seeds and displayed reduced dormancy, indicating that OsFIE1 predominantly functions in late seed development. Ectopic expression of OsFIE1, but not OsFIE2, was deleterious to vegetative growth in a dosage-dependent manner. The newly evolved N-terminal tail of OsFIE1 was probably not the cause of the adverse effects on vegetative growth. The CRISPR/Cas9-derived mutant osfie2 exhibited impaired cellularization of the endosperm, which suggested that OsFIE2 is indispensable for early seed development as a positive regulator of cellularization. Autonomous endosperm was observed in both OsFIE2+- and osfie1/OsFIE2+- but at a very low frequency. Although OsFIE1-PRC2 exhibited H3K27me3 methyltransferase ability in plants, OsFIE1-PRC2 is likely to be less important for development in rice than is OsFIE2-PRC2. Our findings revealed the functional divergence of OsFIE1 and OsFIE2 and shed light on their distinct evolution following duplication.

plant biology↗

bioRxiv2019

Intricate genetic programs controlling dormancy in Mycobacterium tuberculosis

Mycobacterium tuberculosis (MTB), responsible for the deadliest infectious disease worldwide, displays the remarkable ability to transition in and out of dormancy, a hallmark of the pathogens capacity to evade the immune system and opportunistically exploit immunocompromised individuals. Uncovering the gene regulatory programs that underlie the dramatic phenotypic shifts in MTB during disease latency and reactivation has posed an extraordinary challenge. We developed a novel experimental system to precisely control dissolved oxygen levels in MTB cultures in order to capture the chain of transcriptional events that unfold as MTB transitions into and out of hypoxia-induced dormancy. Using a comprehensive genome-wide transcription factor binding location map and insights from network topology analysis, we identified regulatory circuits that deterministically drive sequential transitions across six transcriptionally and functionally distinct states encompassing more than three-fifths of the MTB genome. The architecture of the genetic programs explains the transcriptional dynamics underlying synchronous entry of cells into a dormant state that is primed to infect the host upon encountering favorable conditions.\n\nOne Sentence SummaryHigh-resolution transcriptional time-course reveals six-state genetic program that enables MTB to enter and exit hypoxia-induced dormancy.

systems biology↗

bioRxiv2018

Construction and Characterization of a Synthetic Baculovirus-inducible 39K Promoter

The low expression activity and specificity of natural promoters limit the applications of genetic engineering. To construct a highly efficient synthetic inducible promoter in the Bombyx mori (Lepidoptera), we analyzed the regulatory elements and functional regions of the B. mori nucleopolyhedrovirus (BmNPV) 39K promoter. The results of truncated mutation analysis of the 39K promoter showed that the transcriptional regulatory region spanning positions -573 to -274 and +1 to +62 is essential for virus-inducible promoter activity. Further investigation using electrophoretic mobility shift assay (EMSA) revealed that the baculovirus IE-1 protein binds to the 39K promoter at the -310 to -355 region, and transcription activates the expression of 39K promoter assay. Finally, we successfully constructed a synthetic inducible promoter that increase the virus-inducing activity of other promoters using the baculovirus-inducible transcriptional activation region that binds to specific core elements of 39K (i.e., spanning the region -310 to -355). In summary, we describes a novel, synthetic, and highly efficient biological tool, namely, a virus-inducible 39K promoter, which provides endless possibilities for future gene function research, gene therapy, and pest control in genetic engineering.

bioengineering↗

bioRxiv2018

An essential mycolate remodeling program for mycobacterial adaptation in host cells

The success of Mycobacterium tuberculosis (MTB) stems from its ability to remain hidden from the immune system within macrophages. Here, we report a new technology (Path-seq) to sequence miniscule amounts of MTB transcripts within up to million-fold excess host RNA. Using Path-seq we have discovered a novel transcriptional program for in vivo mycobacterial cell wall remodeling when the pathogen infects alveolar macrophages in mice. We have discovered that MadR transcriptionally modulates two mycolic acid desaturases desA1/A2 to initially promote cell wall remodeling upon in vitro macrophage infection and, subsequently, reduces mycolate biosynthesis upon entering dormancy. We demonstrate that disrupting MadR program is lethal to diverse mycobacteria making this evolutionarily conserved regulator a prime antitubercular target for both early and late stages of infection.\n\nOne Sentence SummaryNovel technology (Path-seq) discovers cell wall remodeling program during Mycobacterium tuberculosis infection of macrophages

microbiology↗