In vivo screening characterizes chromatin factor functions during normal and malignant hematopoiesis

Bulk ex vivo and single-cell in vivo CRISPR knockout screens are used to characterize 680 chromatin factors during mouse hematopoiesis, highlighting lineage-specific and normal and leukemia-specific functions. Cellular differentiation requires extensive alterations in chromatin structure and function, which is elicited by the coordinated action of chromatin and transcription factors. By contrast with transcription factors, the roles of chromatin factors in differentiation have not been systematically characterized. Here, we combine bulk ex vivo and single-cell in vivo CRISPR screens to characterize the role of chromatin factor families in hematopoiesis. We uncover marked lineage specificities for 142 chromatin factors, revealing functional diversity among related chromatin factors (i.e. barrier-to-autointegration factor subcomplexes) as well as shared roles for unrelated repressive complexes that restrain excessive myeloid differentiation. Using epigenetic profiling, we identify functional interactions between lineage-determining transcription factors and several chromatin factors that explain their lineage dependencies. Studying chromatin factor functions in leukemia, we show that leukemia cells engage homeostatic chromatin factor functions to block differentiation, generating specific chromatin factor-transcription factor interactions that might be therapeutically targeted. Together, our work elucidates the lineage-determining properties of chromatin factors across normal and malignant hematopoiesis

LncRNAs: Novel therapeutic targets to treat Multiple Myeloma with RNA-based therapies

Insights into B-cell development and plasma cell biology are essential for understanding the disease known as Multiple Myeloma (MM). The differentiation from a precursor cell into a terminally differentiated, antibody producing plasma cell (PC) is a complex and tightly regulated process, guided by maturation-specific transcriptional programs and external cues such as cytokines present in the bone marrow and secondary lymphoid organs1,2. B lymphocytes develop in the bone marrow (BM) from precursor hematopoietic stem cells (HSC) (Figure 1A), which are the origin of all blood cells3 HSCs differentiate into common lymphoid progenitors that commit to the B-cell lineage due to the expression of lineage-specific transcription factors such as EBF1, PAX5 and E2A3,4. Early BM-dependent stages of B-cell development are structured along the functional rearrangement of immunoglobulin genes, where heavy chain (H- chain) VH-DH-JH segments and light-chain (L-chain) VL-JL segments are rearranged. Finally, early B-cell development is finalized when immature B-cells expressing IgM molecules leave the BM and migrate to secondary lymphoid organs where they further differentiate into Naïve (NBC), follicular or marginal zone B cells1,5. Naïve B cells circulate through peripheral blood and the lymphatic system and enter secondary lymphoid tissues where they can be activated upon exogenous antigen encountering and T cell interaction1,6. Activated B cells can either develop directly into extrafollicular short-lived plasma cells or can mature into germinal center (GC) precursor B-cells6,7. GC microenvironments are formed by proliferating B cells in the follicles of peripheral lymphoid tissues, and are the main site of antibody diversification and affinity maturation (Figure 1B) 6,7. In GCs, B cells are subjected to repeated rounds of somatic hypermutation (SHM) and affinity selection, together with the class-switch recombination (CSR) of heavy chain isotypes, resulting in progressive increase of antibody affinity during immune responses6,7. In brief, SHM is a process that modifies the immunoglobulin variable region (IgV) of the rearranged antibody genes during an immune response, creating a repertoire of diverse B cell receptors whose affinity is “tested” and the best ones are selected. On the other hand, CSR is an irreversible somatic recombination mechanism by which B cells change their immunoglobulin heavy chain expression to gain distinct effector functions6 Finally, clonally selected high-affinity antibody producing GC B cells are terminally differentiated into memory B cells or PCs6,7. Mature PCs are specialized in the production and secretion of high amounts of protective high-affinity antibodies, and they migrate to the BM to turn into long-lived PCs and become central elements of the adaptive immune system1,4. In contrast, the mechanisms that drive memory B cell development are less clear, but they comprise a group of pathogen-experienced cells which are rapidly reactivated upon re- infection4,8

LncRNAs: Novel therapeutic targets to treat Multiple Myeloma with RNA-based therapies

Insights into B-cell development and plasma cell biology are essential for understanding the disease known as Multiple Myeloma (MM). The differentiation from a precursor cell into a terminally differentiated, antibody producing plasma cell (PC) is a complex and tightly regulated process, guided by maturation-specific transcriptional programs and external cues such as cytokines present in the bone marrow and secondary lymphoid organs1,2. B lymphocytes develop in the bone marrow (BM) from precursor hematopoietic stem cells (HSC) (Figure 1A), which are the origin of all blood cells3 HSCs differentiate into common lymphoid progenitors that commit to the B-cell lineage due to the expression of lineage-specific transcription factors such as EBF1, PAX5 and E2A3,4. Early BM-dependent stages of B-cell development are structured along the functional rearrangement of immunoglobulin genes, where heavy chain (H- chain) VH-DH-JH segments and light-chain (L-chain) VL-JL segments are rearranged. Finally, early B-cell development is finalized when immature B-cells expressing IgM molecules leave the BM and migrate to secondary lymphoid organs where they further differentiate into Naïve (NBC), follicular or marginal zone B cells1,5. Naïve B cells circulate through peripheral blood and the lymphatic system and enter secondary lymphoid tissues where they can be activated upon exogenous antigen encountering and T cell interaction1,6. Activated B cells can either develop directly into extrafollicular short-lived plasma cells or can mature into germinal center (GC) precursor B-cells6,7. GC microenvironments are formed by proliferating B cells in the follicles of peripheral lymphoid tissues, and are the main site of antibody diversification and affinity maturation (Figure 1B) 6,7. In GCs, B cells are subjected to repeated rounds of somatic hypermutation (SHM) and affinity selection, together with the class-switch recombination (CSR) of heavy chain isotypes, resulting in progressive increase of antibody affinity during immune responses6,7. In brief, SHM is a process that modifies the immunoglobulin variable region (IgV) of the rearranged antibody genes during an immune response, creating a repertoire of diverse B cell receptors whose affinity is “tested” and the best ones are selected. On the other hand, CSR is an irreversible somatic recombination mechanism by which B cells change their immunoglobulin heavy chain expression to gain distinct effector functions6 Finally, clonally selected high-affinity antibody producing GC B cells are terminally differentiated into memory B cells or PCs6,7. Mature PCs are specialized in the production and secretion of high amounts of protective high-affinity antibodies, and they migrate to the BM to turn into long-lived PCs and become central elements of the adaptive immune system1,4. In contrast, the mechanisms that drive memory B cell development are less clear, but they comprise a group of pathogen-experienced cells which are rapidly reactivated upon re- infection4,8

Deep deconvolution of the haematopoietic stem cell regulatory microenvironment in health, ageing and malignant transformation.

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Loss of the matrix metalloproteinase-10 causes premature features of aging in satellite cells

Aged muscles accumulate satellite cells with a striking decline response to damage. Although intrinsic defects in satellite cells themselves are the major contributors to aging-associated stem cell dysfunction, increasing evidence suggests that changes in the muscle-stem cell local microenvironment also contribute to aging. Here, we demonstrate that loss of the matrix metalloproteinase-10 (MMP-10) in young mice alters the composition of the muscle extracellular matrix (ECM), and specifically disrupts the extracellular matrix of the satellite cell niche. This situation causes premature features of aging in the satellite cells, contributing to their functional decline and a predisposition to enter senescence under proliferative pressure. Similarly, reduction of MMP-10 levels in young satellite cells from wild type animals induces a senescence response, while addition of the protease delays this program. Significantly, the effect of MMP-10 on satellite cell aging can be extended to another context of muscle wasting, muscular dystrophy. Systemic treatment of mdx dystrophic mice with MMP-10 prevents the muscle deterioration phenotype and reduces cellular damage in the satellite cells, which are normally under replicative pressure. Most importantly, MMP-10 conserves its protective effect in the satellite cell-derived myoblasts isolated from a Duchenne muscular dystrophy patient by decreasing the accumulation of damaged DNA. Hence, MMP-10 provides a previously unrecognized therapeutic opportunity to delay satellite cell aging and overcome satellite cell dysfunction in dystrophic muscles

In vivo screening characterizes chromatin factor functions during normal and malignant hematopoiesis

Bulk ex vivo and single-cell in vivo CRISPR knockout screens are used to characterize 680 chromatin factors during mouse hematopoiesis, highlighting lineage-specific and normal and leukemia-specific functions. Cellular differentiation requires extensive alterations in chromatin structure and function, which is elicited by the coordinated action of chromatin and transcription factors. By contrast with transcription factors, the roles of chromatin factors in differentiation have not been systematically characterized. Here, we combine bulk ex vivo and single-cell in vivo CRISPR screens to characterize the role of chromatin factor families in hematopoiesis. We uncover marked lineage specificities for 142 chromatin factors, revealing functional diversity among related chromatin factors (i.e. barrier-to-autointegration factor subcomplexes) as well as shared roles for unrelated repressive complexes that restrain excessive myeloid differentiation. Using epigenetic profiling, we identify functional interactions between lineage-determining transcription factors and several chromatin factors that explain their lineage dependencies. Studying chromatin factor functions in leukemia, we show that leukemia cells engage homeostatic chromatin factor functions to block differentiation, generating specific chromatin factor-transcription factor interactions that might be therapeutically targeted. Together, our work elucidates the lineage-determining properties of chromatin factors across normal and malignant hematopoiesis