Drosophila Sensory Neurons Require Dscam for Dendritic Self-Avoidance and Proper Dendritic Field Organization

SummaryA neuron's dendrites typically do not cross one another. This intrinsic self-avoidance mechanism ensures unambiguous processing of sensory or synaptic inputs. Moreover, some neurons respect the territory of others of the same type, a phenomenon known as tiling. Different types of neurons, however, often have overlapping dendritic fields. We found that Down's syndrome Cell Adhesion Molecule (Dscam) is required for dendritic self-avoidance of all four classes of Drosophila dendritic arborization (da) neurons. However, neighboring mutant class IV da neurons still exhibited tiling, suggesting that self-avoidance and tiling differ in their recognition and repulsion mechanisms. Introducing 1 of the 38,016 Dscam isoforms to da neurons in Dscam mutants was sufficient to significantly restore self-avoidance. Remarkably, expression of a common Dscam isoform in da neurons of different classes prevented their dendrites from sharing the same territory, suggesting that coexistence of dendritic fields of different neuronal classes requires divergent expression of Dscam isoforms

青色発光ダイオードはオプシン3を介し大腸癌細胞のオートファジーを誘導する

Light emitting-diodes (LED) have various effects on living organisms and recent studies have shown the efficacy of visible light irradiation from LED for anticancer therapies. However, the mechanism of LED’s effects on cancer cells remains unclear. The aim of the present study was to investigate the effects of LED on colon cancer cell lines and the role of photoreceptor Opsin 3 (Opn3) on LED irradiation in vitro. Human colon cancer cells (HT-29 or HCT-116) were seeded onto laboratory dishes and irradiated with 465-nm LED at 30 mW/cm2 for 30 minutes. Cell Counting Kit-8 was used to measure cell viability, and apoptosis and caspase 3/8 expression were evaluated by AnnexinV/PI and reverse transcription-polymerase chain reaction (RT-PCR), respectively. Autophagy and expression of LC-3 and beclin-1 were also evaluated by autophagy assays, RT-PCR and Western blotting. We further tested Opn3 knockdown by Opn3 siRNA and the Gi/o G-protein inhibitor NF023 in these assays. Viability of HT-29 and HCT-116 cells was lower in 465-nm LED-irradiated cultures than in control cultures. LC-3 and beclin-1 expressions were significantly higher in LED-irradiated cultures, and autophagosomes were detected in irradiated cells. The reductive effect of cancer cell viability following blue LED irradiation was reversed by Opn3 knockdown or NF023 treatment. Furthermore, increased LC-3 and beclin-1 expression that resulted from blue LED irradiation was suppressed by Opn3 knockdown or NF023 treatment. Blue LED irradiation suppressed the growth of colon cancer cells and Opn3 may play an important role as a photoreceptor

In vivo induction of activin A-producing alveolar macrophages supports the progression of lung cell carcinoma

Alveolar macrophages (AMs) are crucial for maintaining normal lung function. They are abundant in lung cancer tissues, but their pathophysiological significance remains unknown. Here we show, using an orthotopic murine lung cancer model and human carcinoma samples, that AMs support cancer cell proliferation and thus contribute to unfavourable outcome. Inhibin beta A (INHBA) expression is upregulated in AMs under tumor-bearing conditions, leading to the secretion of activin A, a homodimer of INHBA. Accordingly, follistatin, an antagonist of activin A is able to inhibit lung cancer cell proliferation. Single-cell RNA sequence analysis identifies a characteristic subset of AMs specifically induced in the tumor environment that are abundant in INHBA, and distinct from INHBA-expressing AMs in normal lungs. Moreover, postnatal deletion of INHBA/activin A could limit tumor growth in experimental models. Collectively, our findings demonstrate the critical pathological role of activin A-producing AMs in tumorigenesis, and provides means to clearly distinguish them from their healthy counterparts.Taniguchi S., Matsui T., Kimura K., et al. In vivo induction of activin A-producing alveolar macrophages supports the progression of lung cell carcinoma. Nature Communications 14, 143 (2023); https://doi.org/10.1038/s41467-022-35701-8

Single-cell transcriptomics of human cholesteatoma identifies an activin A-producing osteoclastogenic fibroblast subset inducing bone destruction

Cholesteatoma, which potentially results from tympanic membrane retraction, is characterized by intractable local bone erosion and subsequent hearing loss and brain abscess formation. However, the pathophysiological mechanisms underlying bone destruction remain elusive. Here, we performed a single-cell RNA sequencing analysis on human cholesteatoma samples and identify a pathogenic fibroblast subset characterized by abundant expression of inhibin βA. We demonstrate that activin A, a homodimer of inhibin βA, promotes osteoclast differentiation. Furthermore, the deletion of inhibin βA /activin A in these fibroblasts results in decreased osteoclast differentiation in a murine model of cholesteatoma. Moreover, follistatin, an antagonist of activin A, reduces osteoclastogenesis and resultant bone erosion in cholesteatoma. Collectively, these findings indicate that unique activin A-producing fibroblasts present in human cholesteatoma tissues are accountable for bone destruction via the induction of local osteoclastogenesis, suggesting a potential therapeutic target.Shimizu K., Kikuta J., Ohta Y., et al. Single-cell transcriptomics of human cholesteatoma identifies an activin A-producing osteoclastogenic fibroblast subset inducing bone destruction. Nature Communications 14, 4417 (2023); https://doi.org/10.1038/s41467-023-40094-3

Reversible binding and rapid diffusion of proteins in complex with inositol lipids serves to coordinate free movement with spatial information

Polyphosphoinositol lipids convey spatial information partly by their interactions with cellular proteins within defined domains. However, these interactions are prevented when the lipids' head groups are masked by the recruitment of cytosolic effector proteins, whereas these effectors must also have sufficient mobility to maximize functional interactions. To investigate quantitatively how these conflicting functional needs are optimized, we used different fluorescence recovery after photobleaching techniques to investigate inositol lipid–effector protein kinetics in terms of the real-time dissociation from, and diffusion within, the plasma membrane. We find that the protein–lipid complexes retain a relatively rapid (∼0.1–1 µm2/s) diffusion coefficient in the membrane, likely dominated by protein–protein interactions, but the limited time scale (seconds) of these complexes, dictated principally by lipid–protein interactions, limits their range of action to a few microns. Moreover, our data reveal that GAP1IP4BP, a protein that binds PtdIns(4,5)P2 and PtdIns(3,4,5)P3 in vitro with similar affinity, is able to “read” PtdIns(3,4,5)P3 signals in terms of an elongated residence time at the membrane

Protein Kinase Cδ Stimulates Proteasome-Dependent Degradation of C/EBPα during Apoptosis Induction of Leukemic Cells

BACKGROUND:The precise regulation and maintenance of balance between cell proliferation, differentiation and death in metazoan are critical for tissue homeostasis. CCAAT/enhancer-binding protein alpha (C/EBPalpha) has been implicated as a key regulator of differentiation and proliferation in various cell types. Here we investigated the potential dynamic change and role of C/EBPalpha protein during apoptosis induction. METHODOLOGY/PRINCIPAL FINDINGS:Upon onset of apoptosis induced by various kinds of inducers such as NSC606985, etoposide and others, C/EBPalpha expression presented a profound down-regulation in leukemic cell lines and primary cells via induction of protein degradation and inhibition of transcription, as assessed respectively by cycloheximide inhibition test, real-time quantitative RT-PCR and luciferase reporter assay. Applying chemical inhibition, forced expression of dominant negative mutant and catalytic fragment (CF) of protein kinase Cdelta (PKCdelta), which was proteolytically activated during apoptosis induction tested, we showed that the active PKCdelta protein contributed to the increased degradation of C/EBPalpha protein. Three specific proteasome inhibitors antagonized C/EBPalpha degradation during apoptosis induction. More importantly, ectopic expression of PKCdelta-CF stimulated the ubiquitination of C/EBPalpha protein, while the chemical inhibition of PKCdelta action significantly inhibited the enhanced ubiquitination of C/EBPalpha protein under NSC606985 treatment. Additionally, silencing of C/EBPalpha expression by small interfering RNAs enhanced, while inducible expression of C/EBPalpha inhibited NSC606985/etoposide-induced apoptosis in leukemic cells. CONCLUSIONS/SIGNIFICANCE:These observations indicate that the activation of PKCdelta upon apoptosis results in the increased proteasome-dependent degradation of C/EBPalpha, which partially contributes to PKCdelta-mediated apoptosis

Differentiation of human-induced pluripotent stem cell under flow conditions to mature hepatocytes for liver tissue engineering

Hepatic differentiation of human?induced pluripotent stem cells (hiPSCs) under flow conditions in a 3D scaffold is expected to be a major step forward for construction of bioartificial livers. The aims of this study were to induce hepatic differentiation of hiPSCs under perfusion conditions and to perform functional comparisons with fresh human precision?cut liver slices (hPCLS), an excellent benchmark for the human liver in vivo. The majority of the mRNA expression of CYP isoenzymes and transporters and the tested CYP activities, Phase II metabolism, and albumin, urea, and bile acid synthesis in the hiPSC?derived cells reached values that overlap those of hPCLS, which indicates a higher degree of hepatic differentiation than observed until now. Differentiation under flow compared with static conditions had a strong inducing effect on Phase II metabolism and suppressed AFP expression but resulted in slightly lower activity of some of the Phase I metabolism enzymes. Gene expression data indicate that hiPSCs differentiated into both hepatic and biliary directions. In conclusion, the hiPSC differentiated under flow conditions towards hepatocytes express a wide spectrum of liver functions at levels comparable with hPCLS indicating excellent future perspectives for the development of a bioartificial liver system for toxicity testing or as liver support device for patients

Maturation-associated changes in toxicity of the pufferfish Takifugu poecilonotus.

From October 2006 to December 2007, wild specimens of the pufferfish Takifugu poecilonotus (93 females, 45 males) were collected from the Ariake Sea. Tissue toxicity was examined by mouse bioassay, and tetrodotoxin (TTX) content in the blood plasma by enzyme-linked immunosorbent assay. The relationship between toxicity and maturation was investigated based on changes in the gonadosomatic index: December-March in females and November-March in males, the \u27maturation period\u27; April, \u27just after spawning\u27; and the other months, the \u27ordinary period\u27. Toxicity of both sexes was high throughout the year, but sharply declined in April. In all tissues examined (skin, liver, and ovary) other than testis, toxicity exceeded 1000MU/g or 10,000MU/individual in many individuals. Seasonal profiles of tissue toxicity differed markedly between sexes. In females, liver toxicity was high during the ordinary period, and ovary toxicity was high during the maturation period. In males, little maturation-associated change in the toxin distribution was observed. Plasma TTX levels were similar between the sexes (1.59-15.1MU/ml), and fluctuated largely throughout the year without corresponding changes in tissue toxicity. The percentage of TTX binding to high molecular-weight substances in the plasma varied in association with maturation; the binding ratio fluctuated at relatively low levels during the ordinary period, and stabilized at a high level during the maturation period