microPublication biology最新文献

Cross-species expression of proteins in mammalian cells and Xenopus oocytes remains a powerful strategy for functional studies. This can be facilitated by plasmids with dual promoters, enabling use in both systems without separate subcloning. We developed pXeDNA3.1-LacZ, a plasmid that supports expression in mammalian cells and oocytes. The construct combines a cytomegalovirus promoter for mammalian transcription with Xenopus β-globin untranslated regions to enhance oocyte translation, and incorporates XcmI-mediated TA cloning and blue/white selection. To validate it, we used human TRPV1 (HsTRPV1). The vector drove robust expression in HEK293G5A cells and Xenopus oocytes.pXeDNA3.1-LacZ streamlines assays and facilitates studies of membrane proteins.

GABA _A receptors are present in hindbrain and spinal cord networks, playing a pivotal role in regulating locomotion. In this study, we demonstrate that mutations in the gabra4 gene, which encodes the α4 subunit of GABA _A receptors, result in increased swim velocity of larval zebrafish. We also show that this gene is selectively expressed within spinal cord cerebrospinal fluid contacting neurons (CSF-cNs). Given the significance of these neurons in modulating locomotion, our findings support a model in which compromised α4 function leads to an increase in CSF-cN activity, causing a subtle, hyperactive swimming phenotype.

AmiCi24 is a novel siphoviral bacteriophage that was isolated from a soil sample collected in Sewell, NJ, USA using Arthrobacter globiformis B-2979 as the host. AmiCi24 has a genome consisting of 38,466 base pairs that encodes 68 predicted protein-coding genes. Based on gene content, AmiCi24 is assigned to actinobacteriophage cluster AS and subcluster of AS3 and is predicted to be temperate.

Aggressive cell migration is a hallmark of cancer cells. Cancer cells pass through a 3D confined environment during metastasis, which induces mechanical stress on the nucleus. Transwell membranes have been used in research to induce mechanical stress by cell migration in 3D confined space, but other products with microscale pores have not been widely investigated. Here, we report how cancer cells respond to mechanical stress using a TC insert with microscale pores that mimic a 3D confined extracellular environment. As shown in previous studies using the Transwell membranes, our results indicate that the size of microscale pores in the TC insert modulates cancer cell penetration rates and nuclear morphology. Thus, the TC insert is also a useful option to induce mechanical stress by cell migration in a 3D confined environment.

Calcium signaling is an important regulator of stem cell maintenance and differentiation. Here we report the development of an image processing pipeline for ex vivo time-lapse microscopy data that enables the unbiased, automated detection of calcium signaling events in prohemocytes of the Drosophila melanogaster lymph gland. We also show that heterogeneity in gene expression driven by Tep4-Gal4, which is used to mark prohemocytes, accounts for most of the cell-to-cell variability in the signal, and that spontaneous calcium signaling events in the lymph gland can last from a few seconds to well over a minute.

To test the ability of a fission yeast CRISPR-Cas9 system ( SpEDIT ) to carry out genome editing over distance, we constructed a 1,935 bp-long, dsDNA repair template that contained 45 base pair substitutions (SNPs), relative to the wild-type target locus ade6 . Template-directed repair was efficient in the vicinity of the recombination-initiating dsDNA break, but the efficiency fell rapidly with distance (median editing tract length of 163 bp). The regularly distributed markers also revealed evidence for heteroduplex DNA at the ends of repair tracks and, unexpectedly, that DNA ends of the repair template participate in many (~18%) of the genome editing events.

Early growth response 2 (Egr2) is a pleiotropic zinc finger transcription factor with established roles in neural and immune system, but its uterine function remains poorly understood. We found that uterine EGR2 is expressed in luminal epithelium, glandular epithelium, and stroma of human and mouse uteri, with dynamic regulation across the menstrual cycle and early pregnancy. EGR2 expression declined in aged and Sirt1 -deficient mouse uteri, models of reproductive aging. Conditional uterine deletion of EGR2 caused mild subfertility, with fewer litters and total pups per female. These findings indicate EGR2 supports, but is not essential for, uterine function.

After ejaculation, the intrauterine environment undergoes dynamic fluid changes due to post-ejaculated uterine fluid (eUF) coagulation and subsequent liquefaction. These changes presumably contribute to fertilization and reproductive efficiency; however, their physiological roles remain unclear. We studied the significance of the post-ejaculated intrauterine environment during in vivo fertilization. eUF coagulated immediately after ejaculation, and histological analysis of the uterus suggested that eUF liquefaction was promoted 6-10 h post-ejaculation. However, most gametes completed fertilization within 4 h post-ejaculation. Since eUF fluid changes did not align with fertilization timing, they are assumed to contribute to reproductive phenomena beyond sperm transport and release.

The promise of adeno-associated virus (AAV) vectors for gene therapy is held back by the cost and toxicity of the large doses required. This experiment explored the chronopharmacology of AAVs in mice by studying how circadian phase at the time of injection impacted AAV efficacy and revealed that intraperitoneal doses of AAVs injected during the resting phase (ZT6, 12:00PM) produced greater transgene expression over several weeks than equivalent doses injected during the waking phase (ZT18, 12:00AM). This insight could lead to future work that improves safety and reduces costs of AAV gene therapy and other nanoparticle therapies.

In our previous work, we reported a global landscape of opposite aging effects among mouse cell subsets, where each cell subset is defined as a combination of tissue and cell type, and aging leads to increased gene expression in one subset but reduced expression in another. In this study, we investigated whether opposite aging effects are also observed in human cell subsets using the database of differentially expressed genes (DEGs) and differentially accessible regions (DARs) in various human cell subsets. The results suggest that the opposite aging effects occur among human cell subsets at both the transcriptomic and epigenomic levels.