19 research outputs found
Extracellular vesicles from mouse trophoblast cells: effects on neural progenitor cells and potential participants in the placenta-brain Axis
No full textThe fetal brain of the mouse is thought to be dependent upon the placenta as a source of serotonin (5-hydroxytryptamine; 5-HT) and other factors. How factors reach the developing brain remains uncertain but are postulated here to be part of the cargo carried by placental extracellular vesicles (EV). We have analyzed the protein, catecholamine, and small RNA content of EV from mouse trophoblast stem cells (TSC) and TSC differentiated into parietal trophoblast giant cells (pTGC), potential primary purveyors of 5-HT. We have examined how exposure of mouse neural progenitor cells (NPC) to EV from either TSC or pTGC affect their transcriptome profiles. The EV from TB cells contained relatively high amounts of 5-HT, as well as dopamine and norepinephrine, but there were no significant differences between EV derived from pTGC and from TSC. Content of miRNA and small nucleolar (sno)RNA, however, did differ according to EV source, and snoRNA were upregulated in EV from pTGC. The primary inferred targets of the miRNA from both pTGC and TSC were mRNA enriched in the fetal brain. NPC readily internalized EV, leading to changes in their transcriptome profiles. Transcripts regulated were mainly ones enriched in neural tissues. The transcripts in EV-treated NPC that demonstrated a likely complementarity with miRNA in EV were mainly up- rather than down-regulated, with functions linked to neuronal processes. Our results are consistent with placenta-derived EV providing direct support for fetal brain development and being an integral part of the placenta-brain axis.This is a manuscript of an article published as Jessica A Kinkade, Arun S Seetharam, Shrikesh Sachdev, Nathan J Bivens, Brett S Phinney, Gabriela Grigorean, R Michael Roberts, Geetu Tuteja, Cheryl S Rosenfeld, Extracellular vesicles from mouse trophoblast cells: effects on neural progenitor cells and potential participants in the placenta-brain Axis, Biology of Reproduction, 2023;, ioad146, https://doi.org/10.1093/biolre/ioad146. © The Author(s) 2023. Posted with permission
Nuclear Localization of Ikappa Balpha Is Mediated by the Second Ankyrin Repeat: the Ikappa Balpha Ankyrin Repeats Define a Novel Class of cis-Acting Nuclear Import Sequences
No full textThe ability of the Ikappa Balpha protein to sequester dimeric NF-kappa B/Rel proteins in the cytoplasm provides an effective mechanism for regulating the potent transcriptional activation properties of NF-kappa B/Rel family members. Ikappa Balpha can also act in the nucleus as a postinduction repressor of NF-kappa B/Rel proteins. The mechanism by which Ikappa Balpha enters the nucleus is not known, as Ikappa Balpha lacks a discernible classical nuclear localization sequence (NLS). We now report that nuclear localization of Ikappa Balpha is mediated by a novel nuclear import sequence within the second ankyrin repeat. Deletion of the second ankyrin repeat or alanine substitution of hydrophobic residues within the second ankyrin repeat disrupts nuclear localization of Ikappa Balpha . Furthermore, a region encompassing the second ankyrin repeat of Ikappa Balpha is able to function as a discrete nuclear import sequence. The presence of a discrete nuclear import sequence in Ikappa Balpha suggests that cytoplasmic sequestration of the NF-kappa B/Rel-Ikappa Balpha complex is a consequence of the mutual masking of the NLS within NF-kappa B/Rel proteins and the import sequence within Ikappa Balpha . Nuclear import may be a conserved property of ankyrin repeat domains (ARDs), as the ARDs from two other ARD-containing proteins, 53BP2 and GABPbeta , are also able to function as nuclear import sequences. We propose that the Ikappa Balpha ankyrin repeats define a novel class of cis-acting nuclear import sequences
Kelch-like homologue 9 mutation is associated with an early onset autosomal dominant distal myopathy
No full textDistal myopathies are a heterogeneous group of disorders characterized by progressive weakness and muscular atrophy, beginning in distal limb muscles and affecting proximal limb muscles at a later stage. We studied a large German kindred with 10 affected members. Weakness and atrophy of the anterior tibial muscles started between the ages of 8 and 16 years, followed by atrophy of intrinsic hand muscles. Progression was slow, and patients retained the ability to walk until the seventh decade. Serum creatinine kinase levels were increased in the range of 150-1400 U/l. Muscle biopsies showed myopathic changes, whereas immunohistochemistry showed normal expression of marker proteins for muscular dystrophies. Patients had reduced sensation with stocking-glove distribution in the distal limbs in later life. Nerve conduction studies revealed no evidence of neuropathy. Genome-wide linkage analysis in this family revealed a new locus for distal myopathy at 9p21.2-p22.3 (multipoint logarithm of the odds ratio = 4.21). By positional cloning we found a heterozygous mutation L95F in the Kelch-like homologue 9 gene, encoding a bric-a-brac Kelch protein. Molecular modelling indicated that the mutation may interfere with the interaction of the bric-a-brac domain with Cullin 3. Coimmunoprecipitation experiments confirmed that the mutation reduces association with Cullin 3 in the Kelch-like homologue 9-Cullin 3-E3 ubiquitin ligase complex, which is involved in ubiquitin-dependent protein degradation. We identified a unique form of early onset autosomal dominant distal myopathy which is associated with a Kelch-like homologue 9 mutation and interferes with normal skeletal muscle through a novel pathogenetic mechanism.National Institute of Health [AT003389]; Muscular Dystrophy Campaig
Family A and B DNA Polymerases in Cancer: Opportunities for Therapeutic Interventions
No full textDNA polymerases are essential for genome replication, DNA repair and translesion DNA synthesis (TLS). Broadly, these enzymes belong to two groups: replicative and non-replicative DNA polymerases. A considerable body of data suggests that both groups of DNA polymerases are associated with cancer. Many mutations in cancer cells are either the result of error-prone DNA synthesis by non-replicative polymerases, or the inability of replicative DNA polymerases to proofread mismatched nucleotides due to mutations in 3′-5′ exonuclease activity. Moreover, non-replicative, TLS-capable DNA polymerases can negatively impact cancer treatment by synthesizing DNA past lesions generated from treatments such as cisplatin, oxaliplatin, etoposide, bleomycin, and radiotherapy. Hence, the inhibition of DNA polymerases in tumor cells has the potential to enhance treatment outcomes. Here, we review the association of DNA polymerases in cancer from the A and B families, which participate in lesion bypass, and conduct gene replication. We also discuss possible therapeutic interventions that could be used to maneuver the role of these enzymes in tumorigenesis
Continued Complexity of Mutations in Omicron Sublineages
No full textThe latest SARS-CoV-2 variant of concern (VOC), Omicron (B.1.1.529), has diversified into more than 300 sublineages. With an expanding number of newly emerging sublineages, the mutation profile is also becoming complicated. There exist mutually exclusive and revertant mutations in different sublineages. Omicron sublineages share some common mutations with previous VOCs (Alpha, Beta, Gamma, and Delta), indicating an evolutionary relationship between these VOCs. A diverse mutation profile at the spike–antibody interface, flexibility of the regions harboring mutations, mutation types, and coexisting mutations suggest that SARS-CoV-2’s evolution is far from over
Confirmation of murine GBX2 by Western blot and mass spectral analysis.
No full text<p>(A) Western analysis of recombinant GBX2 proteins. The amino acid sequences for GBX2 (B) and GBX2Δ<i>HD</i> (B') recombinant proteins. Bold type indicates matched peptides identified by mass spectrometry. (C) MS/MS fragmentation data table includes: precursor mass (peptide chosen for MS/MS), approximate weight of the band analyzed by mass spectrometry, peptide sequence, location of the peptide in the protein sequence, the Mascot ion score, and the mass error for each peptide sequence. The low mass error score in parts per million (ppm = {[observed mass – theoretical mass]/theoretical mass}×10<sup>6</sup>) suggests that the observed mass matches the theoretical mass.</p
Binding to <i>ROBO1</i> and regulation of NC cell patterning by GBX2.
No full text<p>(A) Gel-shift analysis for identified GBX2 target <i>ROBO1</i>. A reduction in the mobility of [ÿ -32P] ATP labeled <i>ROBO1</i> 100-mer probe is observed with the addition of GBX2 (black arrows), whereas no shift is observed with the addition of GBX2ΔHD (compare lane 2 to lane 4). A supershift is observed in lane 3 with the addition of anti-GBX2. Addition of identical <i>ROBO1</i> 100-mer unlabeled specific competitor probe at 100x, 300x, and 500x molar concentrations in lanes 5–7. Addition of <i>ROBO1</i> 45-mer unlabeled non-specific competitor probe, omitting the GBX2 DNA-binding sequence in lanes 8–10. (B–I) Whole mount in situ hybridization for <i>Robo1</i> (B–E) and the migrating neural crest marker, <i>Sox10</i> (F–I) at gestational stage E9.5, demonstrates abnormal expression in <i>Gbx2<sup>−/−</sup></i> mutants. Image analysis of embryos in a right lateral view (B,C) and dorsal view (D,E) reveals a reduction in expression of <i>Robo1</i> in rhombomere 4 (compare white arrows in B and D to C and E) and disorganized expression in the rhombomere 1 domain (compare black arrows in B and D to C and E) in <i>Gbx2</i><sup>–/–</sup> mutants compared to the WT control. Lateral and dorsal views reveal a reduction in <i>Sox10</i> expression within the otic vesicle (compare F and H to G and I). Two distinct streams of NCCs into pharyngeal arch 1 and pharyngeal arch 2 are defined within control embryos (F, H) whereas in <i>Gbx2</i><sup>−/−</sup> mutants (G,I) the NC streams appear disrupted. In the mutant, expression of <i>Sox10</i> within the NC stream into presumptive pharyngeal arch 1 is significantly downregulated (compare black arrows in F and H to G and I), and the NCCs within the pharyngeal arch 2 appear to be compacted more posteriorly, truncating the stream (compare white arrows in F and H to G and I) when compared to the WT control. ov = otic vesicle.</p
