Sensory deficit screen identifies nsf mutation that differentially affects SNARE recycling and quality control.
2023; 42 (4): 112345
The AAA+ NSF complex is responsible for SNARE complex disassembly both before and after membrane fusion. Loss of NSF function results in pronounced developmental and degenerative defects. In a genetic screen for sensory deficits in zebrafish, we identified a mutation in nsf, I209N, that impairs hearing and balance in a dosage-dependent manner without accompanying defects in motility, myelination, and innervation. Invitro experiments demonstrate that while the I209N NSF protein recognizes SNARE complexes, the effects on disassembly are dependent upon the type of SNARE complex and I209N concentration. Higher levels of I209N protein produce a modest decrease in binary (syntaxin-SNAP-25) SNARE complex disassembly and residual ternary (syntaxin-1A-SNAP-25-synaptobrevin-2) disassembly, whereas at lower concentrations binary disassembly activity is strongly reduced and ternary disassembly activity is absent. Our study suggests that the differential effect on disassembly of SNARE complexes leads to selective effects on NSF-mediated membrane trafficking and auditory/vestibular function.
View details for DOI 10.1016/j.celrep.2023.112345
View details for PubMedID 37027300
Temporal Vestibular Deficits in synaptojanin 1 (synj1) Mutants.
Frontiers in molecular neuroscience
2020; 13: 604189
The lipid phosphatase synaptojanin 1 (synj1) is required for the disassembly of clathrin coats on endocytic compartments. In neurons such activity is necessary for the recycling of endocytosed membrane into synaptic vesicles. Mutations in zebrafish synj1 have been shown to disrupt the activity of ribbon synapses in sensory hair cells. After prolonged mechanical stimulation of hair cells, both phase locking of afferent nerve activity and the recovery of spontaneous release of synaptic vesicles are diminished in synj1 mutants. Presumably as a behavioral consequence of these synaptic deficits, synj1 mutants are unable to maintain an upright posture. To probe vestibular function with respect to postural control in synj1 mutants, we developed a method for assessing the vestibulospinal reflex (VSR) in larvae. We elicited the VSR by rotating the head and recorded tail movements. As expected, the VSR is completely absent in pcdh15a and lhfpl5a mutants that lack inner ear function. Conversely, lhfpl5b mutants, which have a selective loss of function of the lateral line organ, have normal VSRs, suggesting that the hair cells of this organ do not contribute to this reflex. In contrast to mechanotransduction mutants, the synj1 mutant produces normal tail movements during the initial cycles of rotation of the head. Both the amplitude and temporal aspects of the response are unchanged. However, after several rotations, the VSR in synj1 mutants was strongly diminished or absent. Mutant synj1 larvae are able to recover, but the time required for the reappearance of the VSR after prolonged stimulation is dramatically increased in synj1 mutants. Collectively, the data demonstrate a behavioral correlate of the synaptic defects caused by the loss of synj1 function. Our results suggest that defects in synaptic vesicle recycling give rise to fatigue of ribbons synapses and possibly other synapses of the VS circuit, leading to the loss of postural control.
View details for DOI 10.3389/fnmol.2020.604189
View details for PubMedID 33584199
Similarity in gene-regulatory networks suggests that cancer cells share characteristics of embryonic neural cells
JOURNAL OF BIOLOGICAL CHEMISTRY
2017; 292 (31): 12842–59
Cancer cells are immature cells resulting from cellular reprogramming by gene misregulation, and redifferentiation is expected to reduce malignancy. It is unclear, however, whether cancer cells can undergo terminal differentiation. Here, we show that inhibition of the epigenetic modification enzyme enhancer of zeste homolog 2 (EZH2), histone deacetylases 1 and 3 (HDAC1 and -3), lysine demethylase 1A (LSD1), or DNA methyltransferase 1 (DNMT1), which all promote cancer development and progression, leads to postmitotic neuron-like differentiation with loss of malignant features in distinct solid cancer cell lines. The regulatory effect of these enzymes in neuronal differentiation resided in their intrinsic activity in embryonic neural precursor/progenitor cells. We further found that a major part of pan-cancer-promoting genes and the signal transducers of the pan-cancer-promoting signaling pathways, including the epithelial-to-mesenchymal transition (EMT) mesenchymal marker genes, display neural specific expression during embryonic neurulation. In contrast, many tumor suppressor genes, including the EMT epithelial marker gene that encodes cadherin 1 (CDH1), exhibited non-neural or no expression. This correlation indicated that cancer cells and embryonic neural cells share a regulatory network, mediating both tumorigenesis and neural development. This observed similarity in regulatory mechanisms suggests that cancer cells might share characteristics of embryonic neural cells.
View details for DOI 10.1074/jbc.M117.785865
View details for Web of Science ID 000407220100015
View details for PubMedID 28634230
View details for PubMedCentralID PMC5546026
Kruppel-like factor family genes are expressed during Xenopus embryogenesis and involved in germ layer formation and body axis patterning
2015; 244 (10): 1328–46
Kruppel-like factors (Klfs) are a family of transcription factors consisting of 17 members in mammals, Klf1-Klf17, which are involved in fundamental cellular physiological procedures, such as cell proliferation, differentiation, and apoptosis. However, their functions in embryonic development have been poorly understood. Our previous study has demonstrated that the pluripotency factor Klf4 participates in germ layer formation and axis patterning of Xenopus embryos by means of the regulation of key developmental signals. In the present study, we further investigated comprehensively the expression and functions of the klf family genes, klf2, klf5, klf6, klf7, klf8, klf11, klf15, and klf17, during the embryogenesis of Xenopus laevis.Spatio-temporal expression analyses demonstrate that these genes are transcribed both maternally and zygotically in Xenopus embryos, and during organogenesis and tissue differentiation, they are localized to a variety of placodes and tissues. Gain and loss of function studies manifest that Klf factors play different roles in germ layer formation and body axis patterning. Moreover, each Klf factor exhibits distinct regulatory effects on the expression of genes that are essential for germ layer formation and body axis patterning.These results suggest that Klf factors are involved in the fine-tuning of these genes during early embryogenesis.
View details for DOI 10.1002/dvdy.24310
View details for Web of Science ID 000362089300013
View details for PubMedID 26198170
JmjC Domain-containing Protein 6 (Jmjd6) Derepresses the Transcriptional Repressor Transcription Factor 7-like 1 (Tcf7l1) and Is Required for Body Axis Patterning during Xenopus Embryogenesis
JOURNAL OF BIOLOGICAL CHEMISTRY
2015; 290 (33): 20273–83
Tcf7l1 (also known as Tcf3) is a bimodal transcription factor that plays essential roles in embryogenesis and embryonic and adult stem cells. On one hand, Tcf7l1 works as transcriptional repressor via the recruitment of Groucho-related transcriptional corepressors to repress the transcription of Wnt target genes, and, on the other hand, it activates Wnt target genes when Wnt-activated β-catenin interacts with it. However, how its activity is modulated is not well understood. Here we demonstrate that a JmjC-domain containing protein, Jmjd6, interacts with Tcf7l and derepresses Tcf7l. We show that Jmjd6 binds to a region of Tcf7l1 that is also responsible for Groucho interaction, therefore making it possible that Jmjd6 binding displaces the Groucho transcriptional corepressor from Tcf7l1. Moreover, we show that Jmjd6 antagonizes the repression effect of Tcf7l1 on target gene transcription and is able to enhance β-catenin-induced gene activation and that, vice versa, inhibition of Jmjd6 activity compromises gene activation in both cells and Xenopus early embryos. We also show that jmjd6 is both maternally and zygotically transcribed during Xenopus embryogenesis. Loss of Jmjd6 function causes defects in anterioposterior body axis formation and down-regulation of genes that are involved in anterioposterior axis patterning. The results elucidate a novel mechanism underlying the regulation of Tcf7l1 activity and the regulation of embryonic body axis formation.
View details for DOI 10.1074/jbc.M115.646554
View details for Web of Science ID 000359608900027
View details for PubMedID 26157142
View details for PubMedCentralID PMC4536435
Kdm2a/b Lysine Demethylases Regulate Canonical Wnt Signaling by Modulating the Stability of Nuclear beta-Catenin
2015; 33 (6): 660–74
In the absence of Wnt activation, cytosolic β-catenin is degraded through GSK3/CK1-mediated phosphorylation at the N terminus. Here, we show that, upon Wnt activation, the stability of nuclear β-catenin is regulated via methylation/demethylation. The protein lysine demethylases Kdm2a and Kdm2b regulate the turnover of non-phosphorylated β-catenin specifically within the nucleus via direct interaction with the fourth and fifth armadillo repeats. The lysine residues within this region are required for the methylation of non-phosphorylated β-catenin, which is demethylated by Kdm2a/b and subsequently ubiquitylated. During Xenopus embryogenesis, kdm2a/b genes are transcribed during early embryogenesis and are required for the specification of the body axis. Kdm2a/b knockdown in Xenopus embryos leads to increases in non-phosphorylated and methylated β-catenin, concurrent with the upregulation of β-catenin target genes. This mechanism is required for controlling the output of the Wnt/β-catenin signaling pathway to maintain normal cellular functions.
View details for DOI 10.1016/j.devcel.2015.04.006
View details for Web of Science ID 000356773900007
View details for PubMedID 26004508
- Kdm2a/b Lysine Demethylases Regulate Canonical Wnt Signaling by Modulating the Stability of Nuclear β-Catenin Developmental Cell 2015
Klf4 is required for germ-layer differentiation and body axis patterning during Xenopus embryogenesis
2012; 139 (21): 3950–61
Klf4 is a transcription factor of the family of Kruppel-like factors and plays important roles in stem cell biology; however, its function during embryogenesis is unknown. Here, we report the characterization of a Klf4 homologue in Xenopus laevis during embryogenesis. Klf4 is transcribed both maternally and zygotically and the transcript is ubiquitous in embryos during germ-layer formation. Klf4 promotes endoderm differentiation in both Nodal/Activin-dependent and -independent manners. Moreover, Klf4 regulates anteroposterior body axis patterning via activation of a subset of genes in the Spemann organizer, such as Noggin, Dkk1 and Cerberus, which encode Nodal, Wnt and BMP antagonists. Loss of Klf4 function leads to the failure of germ-layer differentiation, the loss of responsiveness of early embryonic cells to inducing signals, e.g. Nodal/Activin, and the loss of transcription of genes involved in axis patterning. We conclude that Klf4 is required for germ-layer differentiation and body axis patterning by means of rendering early embryonic cells competent to differentiation signals.
View details for DOI 10.1242/dev.082024
View details for Web of Science ID 000309701300007
View details for PubMedID 22992953