β-Tubulin Isotype, TUBB4B, Regulates The Maintenance of Cancer Stem Cells
Recent advancements in cancer research have shown that cancer stem cell (CSC) niche is a crucial factor modulating tumor progression and treatment outcomes. It sustains CSCs by orchestrated regulation of several cytokines, growth factors, and signaling pathways. Although the features defining adult stem cell niches are well-explored, the CSC niche is poorly characterized. Since membrane trafficking proteins have been shown to be essential for the localization of critical proteins supporting CSCs, we investigated the role of TUBB4B, a probable membrane trafficking protein that was found to be overexpressed in the membranes of stem cell enriched cultures, in sustaining CSCs in oral cancer. Here, we show that the knockdown of TUBB4B downregulates the expression of pluripotency markers, depletes ALDH1A1+ population, decreases in vitro sphere formation, and diminishes the tumor initiation potential in vivo. As TUBB4B is not known to have any role in transcriptional regulation nor cell signaling, we suspected that its membrane trafficking function plays a role in constituting a CSC niche.
The pattern of its expression in tissue sections, forming a gradient in and around the CSCs, reinforced the notion. Later, we explored its possible cooperation with a signaling protein, Ephrin-B1, the abrogation of which reduces the self-renewal of oral cancer stem cells. Expression and survival analyses based on the TCGA dataset of head and neck squamous cell carcinoma (HNSCC) samples indicated that the functional cooperation of TUBB4 and EFNB1 results in a poor prognosis. We also show that TUBB4B and Ephrin-B1 cohabit in the CSC niche. Moreover, depletion of TUBB4B downregulates the membrane expression of Ephrin-B1 and reduces the CSC population. Our results imply that the dynamics of TUBB4B is decisive for the surface localization of proteins, like Ephrin-B1, that sustain CSCs by their concerted signaling.
IgA Plasma Cells Are Long-Lived Residents of Gut and Bone Marrow That Express Isotype– and Tissue-Specific Gene Expression Patterns
- Antibody secreting plasma cells are made in response to a variety of pathogenic and commensal microbes. While all plasma cells express a core gene transcription program that allows them to secrete large quantities of immunoglobulin, unique transcriptional profiles are linked to plasma cells expressing different antibody isotypes. IgA expressing plasma cells are generally thought of as short-lived in mucosal tissues and they have been understudied in systemic sites like the bone marrow.
- We find that IgA+ plasma cells in both the small intestine lamina propria and the bone marrow are long-lived and transcriptionally related compared to IgG and IgM expressing bone marrow plasma cells. IgA+ plasma cells show signs of shared clonality between the gut and bone marrow, but they do not recirculate at a significant rate and are found within bone marrow plasma cells niches.
- These data suggest that systemic and mucosal IgA+ plasma cells are from a common source, but they do not migrate between tissues. However, comparison of the plasma cells from the small intestine lamina propria to the bone marrow demonstrate a tissue specific gene transcription program. Understanding how these tissue specific gene networks are regulated in plasma cells could lead to increased understanding of the induction of mucosal versus systemic antibody responses and improve vaccine design
Can anti-PGL-I antibody isotypes differentiate leprosy contacts and leprosy patients?
Background: Serological tests for antibody measurement in leprosy have a series of limitations in discriminating contacts and patients. The present paper intends to evaluate if association of more than one antibody isotype in serum samples may be a useful tool in leprosy diagnosis.
Methods: This study evaluated 395 leprosy contacts and 71 leprosy index cases living in endemic municipalities in Northeastern Brazil. The participants were evaluated according to their anti-phenolic glycolipid antigen-I isotype (PGL-I) profile. Serum anti-PGL-I IgM, IgG, and IgA were measured by indirect ELISA.
Results: A strong association was found for antibody positivity in MB leprosy index cases. The odds ratios were 6.11 (95% CI 3.08 – 12.16) for IgM, 3.31 (1.66 – 6.61) for IgG, and 16.97 (8.39 – 34.2) for IgA. For IgM associated with one or more isotypes, the OR was 21.0 (95% CI 10.11 – 43.64), and for IgG + IgA, the OR was 17.58 (6.23 – 49.54). The highest diagnostic sensitivity of 76.0% (95% CI 61.8 – 86.9) was observed for IgM, and the lowest value was 24.1% (13.0 – 38.2), which was observed for IgG + IgA isotypes. Regarding presumptive positive predictive values, the lowest value was obtained for IgM at 24.7% (95% CI 18.1 – 32.3), and the highest values were observed for IgM+ one or more isotypes and for IgG + IgA isotype at 60.0% (44.3 – 74.3) and 66.7% (41.0 – 86.7), respectively.
Conclusions: The present work demonstrated that by associating two or more positive antibody isotypes, the risk of facing a real case of leprosy may increase.
Anti-SARS-CoV-2 Immunoglobulin Isotypes, and Neutralization Activity Against Viral Variants, According to BNT162b2-Vaccination and Infection History
Purpose: To compare SARS-CoV-2 antigen-specific antibody production and plasma neutralizing capacity against B.1 wild-type-like strain, and Gamma/P.1 and Delta/B.1.617.2 variants-of-concern, in subjects with different Covid-19 disease and vaccination histories.
Methods: Adult subjects were: 1) Unvaccinated/hospitalized for Covid-19; 2) Covid-19-recovered followed by one BNT162b2 vaccine dose; and 3) Covid-19-naïve/2-dose BNT162b2 vaccinated. Multiplex Luminex® immunoassays measured IgG, IgA, and IgM plasma levels against SARS-CoV-2 receptor-binding domain (RBD), spike-1 (S), and nucleocapsid proteins. Neutralizing activity was determined in Vero E6 cytopathic assays.
Results: Maximum anti-RBD IgG levels were similar in Covid–nd vaccine dosing; both groups had ≈2‑fold higher anti-RBD IgG levels than Unvaccinated/Covid-19 subjects tracked through 2 weeks post-symptom onset. Anti-S IgG expression patterns were similar to RBD within each group, but with lower signal strengths. Viral antigen-specific IgA and IgM levels were more variable than IgG patterns. Anti-nucleocapsid immunoglobulins were not detected in Covid-19-naïve subjects. Neutralizing activity against the B.1 strain, and Gamma/P.1 and Delta/B.1.617.2 variants, was highest in Covid‑19-recovered/single-dose vaccinated subjects; although neutralization against the Delta variant in this group was only 26% compared to B.1 neutralization, absolute anti-Delta titers suggested maintained protection. Neutralizing titers against the Gamma and Delta variants were 33‒77% and 26‒67%, respectively, versus neutralization against the B.1 strain (100%) in the three groups.
Conclusion: These findings support SARS-CoV-2 mRNA vaccine usefulness regardless of Covid-19 history, and confirm remarkable protection provided by a single vaccine dose in people who have recovered from Covid-19.
Monoclonal Anti-Adenovirus (many isotypes) IgG-FITC conjugate |
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| ADV12-FITC | Alpha Diagnostics | 100 ul | 562.8 EUR |
Anti-All Six Isotypes Of Mammalian Actin Monoclonal Antibody |
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| M01263-1 | BosterBio | 100ul | 476.4 EUR |
Goat Anti-Human IgA+IgG+IgM (H+L)-HRP (for good detection of all 3 isotypes) |
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| 10355-HP | Alpha Diagnostics | 0.5 ml | 270 EUR |
Goat Anti-Human IgA+IgG+IgM (H+L) unlabeled (for good detection of all 3 isotypes) |
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| 10355-UL | Alpha Diagnostics | 0.5 mg | 242.4 EUR |
Goat Anti-Mouse IgG+IgM+IgA, unlabeled IgG (for good detection of 3 isotypes) human serum adsorbed) |
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| 40325-UL | Alpha Diagnostics | 0.5 mg | 196.8 EUR |
Goat Anti-Mouse IgG+IgM+IgA-HRP Conjugate (for good detection of 3 isotypes) human serum adsorbed) |
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| 40325-HP | Alpha Diagnostics | 0.5 ml | 270 EUR |
Mouse Monoclonal Anti-Human IgG1+IgG2+IgG3+IgG4-HRP conjugate (formulated for equal reactivity with all 4 IgG isotypes, 100X, Sufficient for 100 ml) |
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| 10121-24-M | Alpha Diagnostics | 0.1 ml | 634.8 EUR |
Custom development of ELISAs for other species or antibody isotypes not listed in the catalog. Custom testing of samples for IgG/IgM/IgA or total (IgG+IgM+IgA) |
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| 000-CUS | Alpha Diagnostics | Custom | 602 EUR |
Rat IgG-PE conjugate (isotype control) (Isotype control) |
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| 20005-PE | Alpha Diagnostics | 25 tests | 242.4 EUR |
Rat IgG-HRP conjugate (isotype control) (Isotype control) |
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| 20005-HP | Alpha Diagnostics | 100 ug | 196.8 EUR |
Rat IgG-FITC conjugate (isotype control) (Isotype control) |
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| 20005-F | Alpha Diagnostics | 100 ug | 196.8 EUR |
Rat IgG-Biotin conjugate (isotype control) (Isotype control) |
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| 20005-B | Alpha Diagnostics | 100 ug | 196.8 EUR |
Rat IgG Isotype Control |
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| 31-AR15 | Fitzgerald | 10 mg | 183 EUR |
PE Rat IgM, κ Isotype Control |
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| DL22349F-20Tests | DL Develop | 20Tests | 158 EUR |
PE Rat IgM, κ Isotype Control |
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| DL22349F-50Tests | DL Develop | 50Tests | 233 EUR |
PE Rat IgM, κ Isotype Control |
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| DL22364F-25ug | DL Develop | 25μg | 137 EUR |
PE Rat IgM, κ Isotype Control |
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| DL22364F-50ug | DL Develop | 50μg | 182 EUR |
PE rat IgM, κ Isotype Control |
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| E16FRCP004K-025U | EnoGene | 25 μg | 281.67 EUR |
Rat IgG Isotype Control |
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| GWB-37FCE4 | GenWay Biotech | 10 mg | Ask for price |
Rat IgG Isotype Control |
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| GWB-4BFFB2 | GenWay Biotech | 0.5 mg | Ask for price |
Rat IgM Isotype Control |
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| GWB-58F247 | GenWay Biotech | 0.1 mg | Ask for price |
Rat IgM Isotype Control |
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| GWB-5EE54A | GenWay Biotech | 0.5 mg | Ask for price |
Rat IgM Isotype Control |
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| GWB-649C16 | GenWay Biotech | 0.5 mg | Ask for price |
Dog IgG Isotype Control |
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| GWB-323C1E | GenWay Biotech | 5 mg | Ask for price |
Rat IgG Isotype Control |
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| GWB-1E8B74 | GenWay Biotech | 0.1 mg | Ask for price |
Rat IgM Isotype Control |
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| GWB-A128B8 | GenWay Biotech | 0.5 mg | Ask for price |
Rat IgM Isotype Control |
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| GWB-A6187D | GenWay Biotech | 0.5 mg | Ask for price |
Rat IgM Isotype Control |
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| GWB-AAC8DF | GenWay Biotech | 0.1 mg | Ask for price |
The Beta-Tubulin Isotype TUBB6 Controls Microtubule and Actin Dynamics in Osteoclasts
Osteoclasts are bone resorbing cells that participate in the maintenance of bone health. Pathological increase in osteoclast activity causes bone loss, eventually resulting in osteoporosis. Actin cytoskeleton of osteoclasts organizes into a belt of podosomes, which sustains the bone resorption apparatus and is maintained by microtubules. Better understanding of the molecular mechanisms regulating osteoclast cytoskeleton is key to understand the mechanisms of bone resorption, in particular to propose new strategies against osteoporosis. We reported recently that β-tubulin isotype TUBB6 is key for cytoskeleton organization in osteoclasts and for bone resorption. Here, using an osteoclast model CRISPR/Cas9 KO for Tubb6, we show that TUBB6 controls both microtubule and actin dynamics in osteoclasts. Osteoclasts KO for Tubb6 have reduced microtubule growth speed with longer growth life time, higher levels of acetylation, and smaller EB1-caps.
On the other hand, lack of TUBB6 increases podosome life time while the belt of podosomes is destabilized. Finally, we performed proteomic analyses of osteoclast microtubule-associated protein enriched fractions. This highlighted ARHGAP10 as a new microtubule-associated protein, which binding to microtubules appears to be negatively regulated by TUBB6. ARHGAP10 is a negative regulator of CDC42 activity, which participates in actin organization in osteoclasts. Our results suggest that TUBB6 plays a key role in the control of microtubule and actin cytoskeleton dynamics in osteoclasts. Moreover, by controlling ARHGAP10 association with microtubules, TUBB6 may participate in the local control of CDC42 activity to ensure efficient bone resorption.