Biology of the Cell最新文献

The crustacean molting cycle is triggered by the elevation of ecdysteroid levels in the hemolymph during late pre-molt. It is known that these animals absorb water through the intestine and gills to promote bodily swelling and rupture of the old exoskeleton. The participation of two membrane proteins responsible for the most uptake of water during the late pre-molt has been shown in the gill and gut cells of the freshwater shrimp Palaemon argentinus: Na⁺/K⁺-ATPase (NKA), which generates an osmoionic gradient, and Aquaporins (AQPs), water channels, which provide higher water permeability. Studies investigating the action of ecdysteroids on these proteins are scarce. Therefore, we investigated 20-hydroxyecdisone (20E) in relation to the regulation of NKA and AQPs in the gill and gut cells of P. argentinus. We exposed primary cultures of both gut and gill cells to 20E. Gill cells treated with hormone in the presence of NKA-blockers, exhibited a reduction in volume, and cells treated with 20E showed a greater expression of NKA than untreated cells. Additionally, gills cells treated with 20E showed an increase in volume (∼60%), which the aquaporin inhibitor (HgCl₂) prevented. The participation of AQPs in the influx of water was corroborated by a greater expression of AQP in cells treated with 20E compared to untreated cells. Gut cells of animals in intermolt exposed to hormone (20E) maintained their initial cell volume. With the addition of HgCl2, these cells showed a reduction in volume similar to cells of animals in pre-molt. Immunocytochemistry showed a high expression of AQP in gut cells treated with 20E. These findings suggest that 20E regulates the expression of AQP and NKA in the late pre-molt, to provide water uptake for molting. This work offers new perspectives concerning the molting hormone, placing it as a crucial part of water uptake for ecdysis.

The CRCI²NA inaugural symposium, a meeting on tumor and immune ecosystems, took place in the vibrant and picturesque city of Nantes. The meeting gathered world-renowned experts in cancer biology and immunology. It showcased the most advanced science on mechanisms driving cellular heterogeneity, plasticity, and signaling in normal and cancer cellular ecosystems, which contribute to cancer development, progression, and therapeutic resistance. Recent developments in cancer immunotherapy and anti-tumor strategies were also discussed to collectively assess new therapeutic vulnerabilities to defeat cancer.

Backgound Information

Toxoplasma gondii has a relict plastid, the apicoplast, to which nuclear-encoded proteins are targeted after synthesis in the cytosol. Proteins exclusively found in the apicoplast use a Golgi-independent route for trafficking, while dually targeted proteins found in both the apicoplast and the mitochondrion use a Golgi-dependent route. For apicoplast targeting, N-terminal signal sequences have been shown to direct the localization of different reporters. In this study, we use chimeric proteins to dissect out the roles of N-terminal sequences and coding sequences in apicoplast localization and the choice of the trafficking route.

Results

We show that when the N-termini of a dually targeted protein, TgTPx1/2, or of an apicoplast protein, TgACP, are fused with the reporter protein, enhanced green fluorescent protein (eGFP) or endogenous proteins, TgSOD2, TgSOD3, TgACP, or TgTPx1/2, the chimeric proteins exhibit flexibility in apicoplast targeting depending on the coding sequences. Further, the chimeras that are localized to the apicoplast use different trafficking pathways depending on the combination of the N-terminal signals and the coding sequences.

Conclusion and Significance

This report shows, for the first time, that in addition to the N-terminal signal sequences, targeting and trafficking signals also reside within the coding sequences of apicoplast proteins.

Background Information

Microvilli are finger-like, straight, and stable cellular protrusions that are filled with F-actin and present a stereotypical length. They are present in a broad range of cell types across the animal tree of life and mediate several fundamental functions, including nutrient absorption, photosensation, and mechanosensation. Therefore, understanding the origin and evolution of microvilli is key to reconstructing the evolution of animal cellular form and function. Here, we review the current state of knowledge on microvilli evolution and perform a bioinformatic survey of the conservation of genes encoding microvillar proteins in animals and their unicellular relatives.

Results

We first present a detailed description of mammalian microvilli based on two well-studied examples, the brush border microvilli of enterocytes and the stereocilia of hair cells. We also survey the broader diversity of microvilli and discuss similarities and differences between microvilli and filopodia. Based on our bioinformatic survey coupled with carefully reconstructed molecular phylogenies, we reconstitute the order of evolutionary appearance of microvillar proteins. We document the stepwise evolutionary assembly of the “molecular microvillar toolkit” with notable bursts of innovation at two key nodes: the last common filozoan ancestor (correlated with the evolution of microvilli distinct from filopodia) and the last common choanozoan ancestor (correlated with the emergence of inter-microvillar adhesions).

Conclusion and Significance

We conclude with a scenario for the evolution of microvilli from filopodia-like ancestral structures in unicellular precursors of animals.

Background information

Extracellular matrix (ECM)-derived hydrogels are frequently used in three-dimensional (3D) cell culture and organoid formation in several tissues. However, in the 3D cultivation of testicular cells, the hyaluronic acid (HA) hydrogel has not received as much attention. This study examined the effects of three distinct composites, including HA-alginate (HA-Alg), HA-alginate-collagen (HA-Alg-Col), and HA-alginate-decellularized ECM (HA-Alg-dECM), on mouse testicular cell culture and in vitro spermatogenesis.

Methods

For the creation of composites, the concentration of biomaterials used was 0.5% HA, 1% alginate, 2.5 mg/mL collagen, and 25 mg/mL dECM derived from the testicles of Rams. After 3D culture of 5 days post-partum (dpp) mouse testicular cells for 14 days, HA-Alg was selected as a superior composite due to the greater number and size of the produced organoids. Then, cell culture was rerun by HA-Alg for 14 days, which was later extended for an additional 28 days. In addition, the 3D culture of 10 dpp mouse testicular cells was used to compare with 5 dpp mice on day 14. The morphology and gene expression were analyzed using appropriate techniques.

Results

On day 14, the HA-Alg hydrogel showed significantly more organoids in terms of size and number than the other two groups (p < 0.05); nevertheless, none of the groups showed the expected signs of testis organoids. Remarkably, on day 14, the histology and immunostaining tests revealed features of hepatocyte-like cells (HLCs) and albumin production as a marker of HLC functionality. Furthermore, the analysis of gene expression verified the significant expression of angiogenesis markers (p < 0.01). After the extended culture to 28 days, 5 dpp testicular cells once more differentiated into erythrocytes and HLCs, while a small number of organoids showed the characteristic of renal cells. Cell culture of 10 dpp mice for 14 days showed a wide range of cell lineages, including renal, glandular, chondrocyte, and hepatocyte-like cells in comparison to the 5 dpp mice.

Conclusion and significance

While the HA-Alg composite did not support spermatogenesis in the 3D culture of mouse testicular cells, it demonstrated an unpredicted potential for promoting the differentiation of neonate mouse testicular cells into HLC, erythrocytes, and other cell lineages.

Background

The scratch assay is commonly used in cell biology to evaluate cell migration; however, it is not a standardized method; it produces highly variable gap dimensions. We design a printable device, comprising a single wounding tool and a guide, and compared the gap produced by our device and the traditional method. The deviceis printable in a standard 3D printer. Cells were seeded on a 24-well plate. After reaching full confluency, a gap was created using the traditional method (scratch assay with a pipette tip), a pipette tip and the guide of the device, or the single wounding tool and the guide. The gaps were observed for up to 48 h under a light microscope and analyzed.

Results

The results show that the traditional method produces irregular and not straight gaps, and had the worst cell migration rates compared to the other groups. The wounding tool produced scrape signs at the well surface.

Conclusion

The guide and pipette tip delivered the best results for the scratch assay.

Significance

The use of the guide and the pipette tip for the scratch assay allows allows to perform reproducible cell migration experiments.

Background information

Arpin, an Arp2/3 inhibitory protein, inhibits lamellipodial protrusions and cell migration. Arpin expression is lost in tumor cells of several cancer types.

Results

Here we analyzed expression levels of Arpin and various markers using Reverse Phase Protein Array (RPPA) in human mammary carcinomas. We found that Arpin protein levels were correlated with those of several DNA damage response markers. Arpin-null cells display enhanced clustering of double stand breaks (DSBs) when cells are treated with a DNA damaging agent, in line with a previously described role of the Arp2/3 complex in promoting DSB clustering for homologous DNA repair (HDR) in the nucleus. Using a specific HDR assay, we further showed that Arpin depletion increased HDR efficiency two-fold through its ability to inactivate the Arp2/3 complex.

Conclusions

Arpin regulates both cell migration in the cytosol and HDR in the nucleus.

Significance

Loss of Arpin expression coordinates enhanced cell migration with up-regulated DNA repair, which is required when DNA damage is induced by active cell migration.

Background information

Microtubules serve as integral components in cellular operations such as cell division, intracellular trafficking, and cellular architecture. Composed of tubulin protein subunits, these hollow tubular structures have been increasingly elucidated through advanced cryo-electron microscopy (Cryo-EM), which has unveiled the presence of microtubule inner proteins (MIPs) within the microtubular lumen.

Results

In the present investigation, we employ a synergistic approach incorporating high-pressure freezing, cryo-focused ion beam milling, and Cryo-electron tomography (Cryo-ET) to interrogate the in situ architecture of microtubules in Caenorhabditis elegans larvae. Our Cryo-ET assessments across neuronal cilia and diverse tissue types consistently demonstrate the formation of annular configurations within the microtubular lumen.

Conclusions

In concert with recently characterized MIPs, our in situ observations within a living organism corroborate the hypothesis that intricate luminal assemblages exist within microtubule scaffolds. These findings necessitate further exploration into the molecular constituents and functional ramifications of these internal microtubular configurations in both cellular physiology and pathophysiology.