IDO1 + Paneth cells promote immune escape of colorectal cancer

Abstract: Tumors have evolved mechanisms to escape anti-tumor immunosurveillance. They limit humoral and cellular immune activities in the stroma and render tumors resistant to immunotherapy. Sensitizing tumor cells to immune attack is an important strategy to revert immunosuppression. However, the underlying mechanisms of immune escape are still poorly understood. Here we discover Indoleamine-2,3-dioxygenase-1 (IDO1)+ Paneth cells in the stem cell niche of intestinal crypts and tumors, which promoted immune escape of colorectal cancer (CRC). Ido1 expression in Paneth cells was strictly Stat1 dependent. Loss of IDO1+ Paneth cells in murine intestinal adenomas with tumor cell-specific Stat1 deletion had profound effects on the intratumoral immune cell composition. Patient samples and TCGA expression data suggested corresponding cells in human colorectal tumors. Thus, our data uncovered an immune escape mechanism of CRC and identify IDO1+ Paneth cells as a target for immunotherapy

IDO1 + Paneth cells promote immune escape of colorectal cancer

Abstract: Tumors have evolved mechanisms to escape anti-tumor immunosurveillance. They limit humoral and cellular immune activities in the stroma and render tumors resistant to immunotherapy. Sensitizing tumor cells to immune attack is an important strategy to revert immunosuppression. However, the underlying mechanisms of immune escape are still poorly understood. Here we discover Indoleamine-2,3-dioxygenase-1 (IDO1)+ Paneth cells in the stem cell niche of intestinal crypts and tumors, which promoted immune escape of colorectal cancer (CRC). Ido1 expression in Paneth cells was strictly Stat1 dependent. Loss of IDO1+ Paneth cells in murine intestinal adenomas with tumor cell-specific Stat1 deletion had profound effects on the intratumoral immune cell composition. Patient samples and TCGA expression data suggested corresponding cells in human colorectal tumors. Thus, our data uncovered an immune escape mechanism of CRC and identify IDO1+ Paneth cells as a target for immunotherapy

HCT116_YY1_hichip5kb-chr5_40-41_ucsc2.txt.gz

YY1 HiChip data<br

longRange

YY1 HiChip<br><br

longRange

HiChi

PREreview of "GLUD1 dictates muscle stem cell differentiation by controlling mitochondrial glutamate levels"

&lt;p&gt;&lt;strong&gt;This Zenodo record is a permanently preserved version of a PREreview. You can view the complete PREreview at &lt;a href="https://prereview.org/reviews/10066121"&gt;https://prereview.org/reviews/10066121&lt;/a&gt;.&lt;/strong&gt;&lt;/p&gt; &lt;p&gt;This review reflects comments and contributions from Marina Schernthanner and Pablo Ranea-Robles. Review synthesized by Pablo Ranea-Robles.&lt;/p&gt;&lt;p&gt;In this study, the authors studied the role of GLUD1 in satellite cells in the muscle. These cells are responsible for dynamic changes in muscular cells upon different signals, such as exercise. It is well known that metabolic changes play an important role in the fate decision of these cells toward proliferation or differentiation. Here, they found that GLUD1 and glutamine anaplerosis are decreased in differentiation, in contrast to what happens during proliferation. Using inducible KO cells and mice, they show how GLUD1 deficiency induces differentiation to myotubes and imbalance fusion of fibers. This phenotype was associated with an accumulation of glutamate only in mitochondria together with decreased levels of mitochondrial aspartate. In consequence, the malate-aspartate shuttle was inhibited, disturbing the NAD/NADH ratio between the cytosolic and mitochondrial compartments. In conclusion, they establish the role of GLUD1 in muscle satellite cells as a brake on differentiation, allowing proper proliferation using glutamine to feed the TCA cycle. Overall, we found this an excellent study, highly relevant, easy to read, and with multiple techniques and models to sustain their conclusions. Therefore, we would like to congratulate the authors on such a high-impact study. Below, a few comments we think could improve the manuscript and its understanding by the broad scientific community.&lt;/p&gt;&lt;p&gt;Major comments:&lt;/p&gt;&lt;ul&gt;&lt;li&gt;&lt;p&gt;We had no major comments&lt;/p&gt;&lt;/li&gt;&lt;/ul&gt;&lt;p&gt;Minor comments:&lt;/p&gt;&lt;ul&gt;&lt;li&gt;&lt;p&gt;Could authors expand on how they know/tested that the custom media they used worked fine on the protocols of differentiation/proliferation? Was it possible to control for the amount of FBS in proliferation vs differentiation media (there appears to be a big difference between 30% and 0.2% FBS) without affecting the maintenance/differentiation of cells? FBS per se contains a number of growth factors, which could influence metabolomic analyses?&lt;/p&gt;&lt;/li&gt;&lt;li&gt;&lt;p&gt;I would recommend adding some implications/impact of these results more on a big picture over muscular function.&lt;/p&gt;&lt;/li&gt;&lt;li&gt;&lt;p&gt;The graphical abstract is excellent. I would just recommend to make the changes in NAD/NADH more clear, and indicate better the status of WT cell (quiescent vs prolif)&lt;/p&gt;&lt;/li&gt;&lt;li&gt;&lt;p&gt;Are there more number of cells after 48h in fig 2?&lt;/p&gt;&lt;/li&gt;&lt;li&gt;&lt;p&gt;What is the ratio Glu/alpha-kg in the cytosol?&lt;/p&gt;&lt;/li&gt;&lt;li&gt;&lt;p&gt;Metabolic changes were done with proliferation media, as I understand it, why did the authors not perform metabolomic analysis with the differentiation media?&lt;/p&gt;&lt;/li&gt;&lt;li&gt;&lt;p&gt;Figure 5D - can the authors show the metabolite levels of the cytosolic fraction as well (maybe in a supplementary figure)? Given that few differences were observed in whole cell lysates, as shown in B, one would expect that mitochondrial and cytosolic metabolite levels display opposite trends that balance each other out, correct?&lt;/p&gt;&lt;/li&gt;&lt;li&gt;&lt;p&gt;Figure 4 - aren't these minimal differences in transcription upon loss of Glud1 surprising, given the strong phenotypic difference in figure 1? What mediates precocious differentiation if not transcriptional changes? Could it be that the authors are dealing with somewhat heterogeneous populations here and thus relative enrichment of MuSC vs differentiated cell populations might not be readily picked up by bulk RNA-seq?&lt;/p&gt;&lt;/li&gt;&lt;li&gt;&lt;p&gt;Figure 3C-D - couldn't a reduction in GFP+ cells in theory also be due to increased cell death of GFP+ PAX7+ MuSC? Have the authors excluded this possibility by f.e. showing that there is no difference in TUNEL+(general cell death, DNA damage marker) and Caspase-3+ (apoptotic) cells between KO and WT MuSCs? For D) addition of nuclear signals to indicate cell fusion as expected in muscle fibers/myotubes would be helpful.&lt;/p&gt;&lt;/li&gt;&lt;/ul&gt;&lt;p&gt;Suggestions for future studies:&lt;/p&gt;&lt;ul&gt;&lt;li&gt;&lt;p&gt; It would be extremely interesting to evaluate a possible rescue of the phenotype in vivo. Perhaps with alanine supplementation?&lt;/p&gt;&lt;/li&gt;&lt;/ul&gt; &lt;h2&gt;Competing interests&lt;/h2&gt; &lt;p&gt; The author declares that they have no competing interests. &lt;/p&gt

PREreview of "A kidney-hypothalamus axis promotes compensatory glucose production in response to glycosuria"

This Zenodo record is a permanently preserved version of a PREreview. You can view the complete PREreview at https://prereview.org/reviews/8431146. This review reflects comments and contributions from Marina Schernthanner, Femi Arogundade and Pablo Ranea-Robles. Review synthesized by Jonny Coates. The study leverages the phenotype presented by the renal Glut2 KO mice (glycosuria with normal glycemia) to investigate how the body senses this glucose loss and the mechanisms behind metabolic homeostasis processes that lead to enhanced glucose production so glycemia remains stable. The use of a genetically modified mouse model with renal Glut2 knockout provides a controlled system for studying the specific role of renal glucose transporters in glucose homeostasis. The study involves various methods, including measurements of glucose production, metabolomics, gene expression related to the hypothalamic-pituitary-adrenal axis, afferent renal nerve ablation, and analysis of secreted proteins. The authors point to a kidney/hypothalamus axis and suggest the involvement of different acute phase proteins in this homeostatic response. The limitations of the study are acknowledged, and further research is suggested to delve deeper into the role of secretory proteins and the specific source of endogenous glucose production after afferent renal denervation. The manuscript is well written and the results are potentially of interest. The study's findings have potential implications for the field of diabetes treatment, as they suggest a mechanism that may explain why SGLT2 inhibitors don't achieve their full potential in lowering blood glucose levels. However, we think that some of the conclusions are merely based on descriptive assessments of changes occurring in the renal Glut2 KO mice. There are a lack of details in the reporting of some of the results and, in particular, in the discussion section, that would also require a bit more explanation from the authors. Right now, it could be hard for the reader to place this research in context. We have summarized our comments below Major comments: The study mentions the use of male and female mice, but it's important to know the sample sizes for each experimental group and how gender might influence the results. Additionally, the authors should provide more details about the control groups and their matching criteria to ensure the validity of comparisons. Moreover, the exact genetic information for the knockout mice i.e. what is the CreER driver that makes it kidney-specific? Is missing. It is currently inconsistent in terms of sex and age of mice used for different experiments. Minor comments: While the metabolomics analysis is described, more information is needed about the biological significance of the changes observed in the metabolites. How do these changes relate to the compensatory glucose production, and are they causally linked? The paper would benefit from improved organization and clarity, particularly in the results and discussion sections. Crh+ cells in control image of fig 2 are not clear. The authors could consider highlighting the are where these cells are present, or add an inset showing a zoomed image of some positive cells How specific is the use of capsaicin to selectively suppress afferent renal nerve activity? Does this impact other neurons? Either citations or experimental data should be included here. The conditions of mice in Sup Fig. 1 are not clear and should be stated clearly in this part of the text and in the figure legend. The study is transparent about its limitations and raises important questions for future research. This acknowledgment of limitations contributes to the scientific rigor of the work. While control groups are mentioned, it's not clear how these controls were chosen or matched to the experimental group. Further information is needed on how these controls were used to make valid comparisons. While the study describes the experimental procedures in detail, it's essential to provide information on how many times these experiments were replicated to assess the reproducibility of the results. This is especially crucial given the complex methods used. Blocking the HPA axis and assessing responses in KO and WT mice would strengthen the data in Fig 2 Investigating or showing the levels of glucagon and adrenaline to delineate mechanisms of tissue-specific glucose production would further strengthen the data presented. Is it possible to measure glucose production under denervation conditions? That would support the conclusion if the increased glucose production is blunted Not everyone might be familiar with the abbreviation 2D-DIGE. Explaining this before first use would be beneficial. Supp fig 2 could be fused with Fig 4 to make the argument more convincing. The authors state that "It is possible that afferent renal denervation in the present study attenuated only hepatic glucose production through the hypothalamus without affecting the compensatory increase in renal (local) glucose production". Addressing this would significantly strengthen the manuscript, particularly given that the title includes "hypothalamus-kidney axis". Comments on reporting: The paper mentions the use of statistical tests but lacks information on the specific statistical tests performed for each analysis. It's crucial to provide details on the tests used, assumptions made, and how p-values were adjusted for multiple comparisons, if applicable. Suggestions for future studies: Extend the research to human subjects, particularly individuals with diabetes treated with SGLT2 inhibitors. Investigate whether similar mechanisms and pathways are at play in humans, and whether these findings have clinical relevance. Investigate the specific roles of secreted proteins, such as acute phase proteins and major urinary proteins, in glucose regulation and potential interactions with the kidney-hypothalamus axis. Explore how the kidney-hypothalamus axis integrates with other nervous system and endocrine signals involved in glucose regulation, such as insulin and glucagon. Conduct in-depth studies on the impact of afferent renal nerve activity on glucose homeostasis and the signaling pathways involved. Investigate the role of sensory nerves in detecting glycosuria and triggering compensatory responses. Competing interests The author declares that they have no competing interests

PREreview of "shRNA drop-out screen identifies BRD4 targeting transcription from RNA polymerase II system to activate β-catenin to promote soft-tissue tumor proliferations"

This Zenodo record is a permanently preserved version of a PREreview. You can view the complete PREreview at https://prereview.org/reviews/8333931. This review reflects comments and contributions from Arpita Ghosh, Teena Bajaj, Rebecca A. Shelley, Marina Schernthanner, Sourav Mukherjee, Emma Phillips & Femi Arogundade. Review synthesized by Arpita Ghosh & Garima Jain. Brief summary of the study The authors present a screen carried out in soft-tissue tumour cells, in which the chromatin reader BRD4 was silenced in the presence of PDGF-BB, which was included to promote proliferation. BRD4 was chosen as it was identified in their lab in a previous screen to be important for soft-tissue tumour cell growth. They explore the effects of BRD4 silencing and inhibition on soft-tissue tumour cell morphology, proliferation and cell cycle. Some settings relate effects of BRD4 silencing to stimulation of the cells using PDGF-BB, a cytokine which, amongst other things, is known to activate beta-catenin signalling. Major comments This preprint was extremely difficult to read as the authors did not clearly explain why or how they performed the experiments and over-state their findings throughout the manuscript. There are major omissions of the reporting of the methodologies behind key experiments - most notably how the screen was performed and what kind of cells were used. In order to improve the community value of this preprint, the authors should carefully revisit it and reframe their data in terms of what it actually concretely shows, removing all statements which are not backed up by their experiments. Any conclusions about molecular interactions, transcriptional activity, tumorigenesis, prognosis etc are not supported by the data in this paper. The authors very often make bold statements, using language such as "strongly suggest," "key regulator" and "driver," throughout. These should be toned down or removed. The authors should also take care to cite any previous findings they are referring to - such as the ChipSeq data they refer to here. The title of this manuscript clearly emphasises the shRNA screen, which is only shown and mentioned in the last figure. In addition, the ordering of the figures is not very intuitive - it might make more sense to start with the findings from figure 6 (i.e. introduce the shRNA screen, which also highlights key proteins) and then delve into mechanistic details as done in figures 1-5 A very thorough proofreading of the entire manuscript is required to make sentences more conducive, comprehensible and error free (tense and grammar). Many sentences are incomplete in their meaning. For example, wherever there is an "effect" mentioned, on what the effect is observed is usually missing, or explanation is missing for what a "tumor D" cells are, etc. Looking at pRB levels is not enough to draw conclusions about apoptosis. The authors should consider exploring caspase activity or annexinV staining to investigate apoptosis. Further, an increase in subG1 DNA levels in the cell cycle analysis assay is expected if cells are dying. Additionally, it would be good to check the total GSK levels. More details are required for the shRNA screen mentioning controls. Top hits should be plotted and lists should be provided for the same as supplementary information. To comment on cellular proliferation as shown on Figure 1A, a Ki67 or BrdU assay would be required. The cell density plays a major role in governing cell shape and area. From the representative images in Figure 2(a), it is evident that cell density of shNTC is higher than BRD4 sh#1 and BRD4 sh#2. To strongly claim the observation from the Figure, better representative images should be chosen and details for cell number stated in the methods. Vinculin expression is decreased upon BRD4 knock down. Why Vinculin was chosen with pRb is unclear. While in the results section, the changes in the Vinculin expression is not mentioned. If Vinculin was chosen as a control, then a better control must be chosen as Vinculin expression depends upon changes in cellular cytoskeleton i.e., cell shape or cell area. In Figure 4, the authors did not distinguish between nuclear and cytoplasmic b-catenin. Given that they look at b-catenin target genes, they should have probably focused on nuclear protein fractions/lysates and then normalised the levels to a nuclear housekeeping gene. In Figure 5, the actin levels at 800 nM are reduced by approximately the same amount as the reduction seen for B-catenin and c-Abl. It would be recommended that the authors repeat the experiment and show quantification and statistics or remove these claims. In the discussion, the tone needs to be dialled down for the overhyped statements and claims which were not tested experimentally. For example, no interaction studies were carried out. Explanation for what "sorafenib" is and why it is relevant to the discussion of the data presented here is missing. More data would be required to make claims about the effect of BRD4 knockdown on the differentiation status of the cells used in this study (eg, levels of differentiation markers). The data do not allow for any conclusions about prognosis. The claim that "transcription from RNA polymerase II" activity is regulated cannot be made by simply showing that some of the candidate genes from the screen are part of this signature. The authors would need to perform additional experiments to test the involvement of RNA polymerase II. The authors showed that BRD4 silencing reduced HIF-1a expression but with that they cannot claim that BRD4 "activates" HIF-1a in these cells. These claims should be removed or should be stated as hypotheses and should definitely not appear in the title. Minor comments All the acronyms used should be first introduced with their full forms and also a line stating why and how that is related to this study would be useful for readers and reviewers. A Figure explaining the methodology for shRNA screen need not be a primary figure as it is widely known in the field. Although this can be incorporated as a supplementary figure. Western blots seem edited with very high contrast, should be kept neutral to observe actual effects, or else raw figures for the blots can be put in the supplementary file. Microscopy images provided for cells have very low visibility. Better images should be provided. Missing figure legends. Comments on reporting Cell lines used should be clearly stated. Methods used are very vague without stating detail like concentrations used for inhibitors or in general in explaining the protocols. Mutations done are very unclear; what mutations, where they have been introduced and also the rationale behind them needs to be stated. All observations reported should be re-checked carefully like "significantly upregulated 72" does not state what exactly 72 is. It is not clear exactly how any statistical measurements were made for the experiments. The figure legend reports n=8. The authors should clarify whether they mean 8 cells were measured or 8 independent experiments. Statistical significance is missing from many figures. All the pathways and upstream or downstream targets mentioned in the manuscript body should be re-visited to elaborate and clearly state what target is upstream and downstream of what other target. Suggestions for future work: Re-organizing the entire manuscript to make the story more interesting and also adding value to the title of the paper. Compiling a few figure panels together, or rearranging their order might be necessary to frame the story better. Competing interests The author declares that they have no competing interests

IDO1 + Paneth cells promote immune escape of colorectal cancer

Abstract: Tumors have evolved mechanisms to escape anti-tumor immunosurveillance. They limit humoral and cellular immune activities in the stroma and render tumors resistant to immunotherapy. Sensitizing tumor cells to immune attack is an important strategy to revert immunosuppression. However, the underlying mechanisms of immune escape are still poorly understood. Here we discover Indoleamine-2,3-dioxygenase-1 (IDO1)+ Paneth cells in the stem cell niche of intestinal crypts and tumors, which promoted immune escape of colorectal cancer (CRC). Ido1 expression in Paneth cells was strictly Stat1 dependent. Loss of IDO1+ Paneth cells in murine intestinal adenomas with tumor cell-specific Stat1 deletion had profound effects on the intratumoral immune cell composition. Patient samples and TCGA expression data suggested corresponding cells in human colorectal tumors. Thus, our data uncovered an immune escape mechanism of CRC and identify IDO1+ Paneth cells as a target for immunotherapy