ULK1 and ULK2 are less redundant than previously thought:computational analysis uncovers distinct regulation and functions of these autophagy induction proteins

Macroautophagy, the degradation of cytoplasmic content by lysosomal fusion, is an evolutionary conserved process promoting homeostasis and intracellular defence. Macroautophagy is initiated primarily by a complex containing ULK1 or ULK2 (two paralogs of the yeast Atg1 protein). To understand the differences between ULK1 and ULK2, we compared the human ULK1 and ULK2 proteins and their regulation. Despite the similarity in their enzymatic domain, we found that ULK1 and ULK2 have major differences in their autophagy-related interactors and their post-translational and transcriptional regulators. We identified 18 ULK1-specific and 7 ULK2-specific protein motifs serving as different interaction interfaces. We found that interactors of ULK1 and ULK2 all have different tissue-specific expressions partially contributing to diverse and ULK-specific interaction networks in various tissues. We identified three ULK1-specific and one ULK2-specific transcription factor binding sites, and eight sites shared by the regulatory region of both genes. Importantly, we found that both their post-translational and transcriptional regulators are involved in distinct biological processes—suggesting separate functions for ULK1 and ULK2. Unravelling differences between ULK1 and ULK2 could lead to a better understanding of how ULK-type specific dysregulation affects autophagy and other cellular processes that have been implicated in diseases such as inflammatory bowel disease and cancer

Hyaluronic acid-GPRC5C signalling promotes dormancy in haematopoietic stem cells

Altres ajuts: The Behrens-Weise-Foundation, the German Research Foundation (DFG) under the German Excellence Strategy (CIBSS-EXC-2189, project ID 390939984), SFB1425 (Project #422681845), SFB992 (Project #192904750; B07), SFB1479 (P05); Wellcome-MRC Cambridge Stem Cell Institute (203151/Z/16/Z)Bone marrow haematopoietic stem cells (HSCs) are vital for lifelong maintenance of healthy haematopoiesis. In inbred mice housed in gnotobiotic facilities, the top of the haematopoietic hierarchy is occupied by dormant HSCs, which reversibly exit quiescence during stress. Whether HSC dormancy exists in humans remains debatable. Here, using single-cell RNA sequencing, we show a continuous landscape of highly purified human bone marrow HSCs displaying varying degrees of dormancy. We identify the orphan receptor GPRC5C, which enriches for dormant human HSCs. GPRC5C is also essential for HSC function, as demonstrated by genetic loss- and gain-of-function analyses. Through structural modelling and biochemical assays, we show that hyaluronic acid, a bone marrow extracellular matrix component, preserves dormancy through GPRC5C. We identify the hyaluronic acid-GPRC5C signalling axis controlling the state of dormancy in mouse and human HSCs

Mouse multipotent progenitor 5 cells are located at the interphase between hematopoietic stem and progenitor cells

International audienceAbstract Hematopoietic stem cells (HSCs) and distinct multipotent progenitor (MPP) populations (MPP1-4) contained within the Lin−Sca-1+c-Kit+ (LSK) compartment have previously been identified using diverse surface-marker panels. Here, we phenotypically define and functionally characterize MPP5 (LSK CD34+CD135−CD48−CD150−). Upon transplantation, MPP5 supports initial emergency myelopoiesis followed by stable contribution to the lymphoid lineage. MPP5, capable of generating MPP1-4 but not HSCs, represents a dynamic and versatile component of the MPP network. To characterize all hematopoietic stem and progenitor cells, we performed RNA-sequencing (RNA-seq) analysis to identify specific transcriptomic landscapes of HSCs and MPP1-5. This was complemented by single-cell RNA-seq analysis of LSK cells to establish the differentiation trajectories from HSCs to MPP1-5. In agreement with functional reconstitution activity, MPP5 is located immediately downstream of HSCs but upstream of the more committed MPP2-4. This study provides a comprehensive analysis of the LSK compartment, focusing on the functional and molecular characteristics of the newly defined MPP5 subset

GPRC5C drives branched-chain amino acid metabolism in leukemogenesis

Altres ajuts: Jose Carreras Leukämie-StiftungLeukemia stem cells (LSCs) share numerous features with healthy hematopoietic stem cells (HSCs). G-protein coupled receptor family C group 5 member C (GPRC5C) is a regulator of HSC dormancy. However, GPRC5C functionality in acute myeloid leukemia (AML) is yet to be determined. Within patient AML cohorts, high GPRC5C levels correlated with poorer survival. Ectopic Gprc5c expression increased AML aggression through the activation of NF-κB, which resulted in an altered metabolic state with increased levels of intracellular branched-chain amino acids (BCAAs). This onco-metabolic profile was reversed upon loss of Gprc5c, which also abrogated the leukemia-initiating potential. Targeting the BCAA transporter SLC7A5 with JPH203 inhibited oxidative phosphorylation and elicited strong antileukemia effects, specifically in mouse and patient AML samples while sparing healthy bone marrow cells. This antileukemia effect was strengthened in the presence of venetoclax and azacitidine. Our results indicate that the GPRC5C-NF-κB-SLC7A5-BCAAs axis is a therapeutic target that can compromise leukemia stem cell function in AML

A Truncated Form of IKKa Is Responsible for Specific Nuclear IKK Activity in Colorectal Cancer

Nuclear IKK alpha regulates gene transcription by phosphorylating specific substrates and has been linked to cancer progression and metastasis. However, the mechanistic connection between tumorigenesis and IKK alpha activity remains poorly understood. We have now analyzed 288 human colorectal cancer samples and found a significant association between the presence of nuclear IKK and malignancy. Importantly, the nucleus of tumor cells contains an active IKK alpha isoform with a predicted molecular weight of 45 kDa (p45-IKK alpha) that includes the kinase domain but lacks several regulatory regions. Active nuclear p45-IKK alpha forms a complex with nonactive IKK alpha and NEMO that mediates phosphorylation of SMRT and histone H3. Proteolytic cleavage of FL-IKK alpha into p45-IKK alpha is required for preventing the apoptosis of CRC cells in vitro and sustaining tumor growth in vivo. Our findings identify a potentially druggable target for treating patients with advance refractory CRC

A Journey Into the Unknown: PhD Students in a European Training Network on Age-related Changes in Hematopoiesis Conduct Their Project During a Global Pandemic

3 p.-1 fig.This project has received funding from the European Union’s Horizon 2020 Research and Innovation Program under the Marie Skłodowska-Curie grant agreement no. 813091 (ARCH, age-related changes in hematopoiesis).Peer reviewe

Multilayer omics analysis reveals a non-classical retinoic acid signaling axis that regulates hematopoietic stem cell identity

Hematopoietic stem cells (HSCs) rely on complex regulatory networks to preserve stemness. Due to the scarcity of HSCs, technical challenges have limited our insights into the interplay between metabolites, transcription, and the epigenome. In this study, we generated low-input metabolomics, transcriptomics, chromatin accessibility, and chromatin immunoprecipitation data , revealing distinct metabolic hubs that are enriched in HSCs and their downstream multipotent progenitors. Mechanistically, we uncover a non-classical retinoic acid (RA) signaling axis that regulates HSC function. We show that HSCs rely on Cyp26b1, an enzyme conventionally considered to limit RA effects in the cell. In contrast to the traditional view, we demonstrate that Cyp26b1 is indispensable for production of the active metabolite 4-oxo-RA. Further, RA receptor beta (Rarb) is required for complete transmission of 4-oxo-RA-mediated signaling to maintain stem cells. Our findings emphasize that a single metabolite controls stem cell fate by instructing epigenetic and transcriptional attributes

Hyaluronic acid-GPRC5C signalling promotes dormancy in haematopoietic stem cells.

Funder: Wellcome TrustBone marrow haematopoietic stem cells (HSCs) are vital for lifelong maintenance of healthy haematopoiesis. In inbred mice housed in gnotobiotic facilities, the top of the haematopoietic hierarchy is occupied by dormant HSCs, which reversibly exit quiescence during stress. Whether HSC dormancy exists in humans remains debatable. Here, using single-cell RNA sequencing, we show a continuous landscape of highly purified human bone marrow HSCs displaying varying degrees of dormancy. We identify the orphan receptor GPRC5C, which enriches for dormant human HSCs. GPRC5C is also essential for HSC function, as demonstrated by genetic loss- and gain-of-function analyses. Through structural modelling and biochemical assays, we show that hyaluronic acid, a bone marrow extracellular matrix component, preserves dormancy through GPRC5C. We identify the hyaluronic acid-GPRC5C signalling axis controlling the state of dormancy in mouse and human HSCs

A Journey Into the Unknown: PhD Students in a European Training Network on Age-related Changes in Hematopoiesis Conduct Their Project During a Global Pandemic

The age-related changes in hematopoiesis (ARCH) project is part of the Innovative Training Network (ITN) of the Marie-Sklodowska Curie Actions (MSCA) program, which provides doctorate training of excellence based on the exchange of ideas and competencies from the academic and private sectors.1 The ARCH project intends to outline hematopoietic stem cell (HSC) alterations with aging, how hematopoietic cell individuality is controlled at the transcriptional and epigenetic levels in normal hematopoiesis and in leukemias, and understand the crosstalk between intrinsic and extrinsic indications that support the proliferation of preleukemic and leukemic cells within the hematopoietic niche. We are 15 PhD students funded by this network, based around Europe, and our common aim is to understand functional changes in the hematopoietic system with age, how these changes link to the development of age-associated diseases and in parallel work towards the development of new treatments.2 Our projects kicked off just when the severe acute respiratory coronavirus-2 (SARS-CoV-2) emerged. Two and a half years later, SARS-CoV-2 continues to infect millions of people and has taken the lives of at least 6 million people worldwide.3 The COVID-19 outbreak brought along social isolation and feelings of uncertainty to everyone around the world, including doctorate students.4,5 Ironically, our projects have been more relevant than ever, as the pandemic has highlighted the important relationship between age-related changes in hematopoiesis and disease severity. Below, we aim to discuss the timeline of the ARCH project throughout the pandemic and how we managed to courageously pull through the hardships of doing research during a global pandemic within different settings (academia/institutes and industry). We provide recommendations to future PhD students on how to manage their PhD projects during global emergencies.The age-related changes in hematopoiesis (ARCH) project is part of the Innovative Training Network (ITN) of the Marie-Sklodowska Curie Actions (MSCA) program, which provides doctorate training of excellence based on the exchange of ideas and competencies from the academic and private sectors.1 The ARCH project intends to outline hematopoietic stem cell (HSC) alterations with aging, how hematopoietic cell individuality is controlled at the transcriptional and epigenetic levels in normal hematopoiesis and in leukemias, and understand the crosstalk between intrinsic and extrinsic indications that support the proliferation of preleukemic and leukemic cells within the hematopoietic niche. We are 15 PhD students funded by this network, based around Europe, and our common aim is to understand functional changes in the hematopoietic system with age, how these changes link to the development of age-associated diseases and in parallel work towards the development of new treatments.2 Our projects kicked off just when the severe acute respiratory coronavirus-2 (SARS-CoV-2) emerged. Two and a half years later, SARS-CoV-2 continues to infect millions of people and has taken the lives of at least 6 million people worldwide.3 The COVID-19 outbreak brought along social isolation and feelings of uncertainty to everyone around the world, including doctorate students.4,5 Ironically, our projects have been more relevant than ever, as the pandemic has highlighted the important relationship between age-related changes in hematopoiesis and disease severity. Below, we aim to discuss the timeline of the ARCH project throughout the pandemic and how we managed to courageously pull through the hardships of doing research during a global pandemic within different settings (academia/institutes and industry). We provide recommendations to future PhD students on how to manage their PhD projects during global emergencies