Periodic Reporting for period 1 - StemLinc (Identification and characterization of long non-coding RNAs as drivers of stemness in hematopoietic stem cells and leukemia.)
Reporting period: 2023-01-01 to 2024-12-31
Hematopoietic stem cells (HSCs) are a rare and essential population responsible for maintaining lifelong blood cell production. Their ability to self-renew and differentiate is crucial for sustaining a healthy blood and immune system. However, disruptions in these regulatory processes can result in hematological disorders, including leukemia. Understanding the molecular mechanisms that govern HSC identity and function is fundamental to uncovering how such dysregulations contribute to disease development.
Long non-coding RNAs (lncRNAs) have emerged as key regulators of gene expression, yet their roles in HSC biology and leukemia remain largely unexplored. Unlike protein-coding RNAs, lncRNAs function as regulatory molecules, influencing gene activity and cellular behavior. Recent evidence suggests that lncRNAs play a crucial role in stem cell function, making them promising candidates for understanding hematopoiesis and developing novel therapeutic approaches for leukemia and other blood disorders.
The StemLinc project, funded by the Marie Skłodowska-Curie Actions (MSCA) program, aimed to identify and characterize lncRNAs involved in the regulation of HSC stemness and their potential role in leukemia.
The project’s specific objectives were:
To generate a comprehensive catalog of hematopoietic lncRNAs through deep total RNA sequencing of primary mouse blood cells and bioinformatics analysis.
To infer the function of identified lncRNAs using computational approaches integrating genomic, epigenomic and transcriptomic data.
To validate the role of selected candidate lncRNAs through functional experiments and assess their relevance in leukemia.
By integrating advanced transcriptomics, functional genomics, and computational analyses, StemLinc significantly advances multiple scientific domains:
Hematopoietic system biology: Providing new insights into the transcriptional landscape of HSCs, progenitors, and differentiated blood cells.
ncRNA biology: Characterizing previously unexplored lncRNAs, including mono-exonic transcripts, broadening the understanding of non-coding RNA regulation in hematopoiesis.
Resource development for future research: Identifying candidate lncRNAs strongly associated with HSC identity and function, offering valuable insights for disease biomarker discovery and future large-scale studies.
Long non-coding RNAs (lncRNAs) have emerged as key regulators of gene expression, yet their roles in HSC biology and leukemia remain largely unexplored. Unlike protein-coding RNAs, lncRNAs function as regulatory molecules, influencing gene activity and cellular behavior. Recent evidence suggests that lncRNAs play a crucial role in stem cell function, making them promising candidates for understanding hematopoiesis and developing novel therapeutic approaches for leukemia and other blood disorders.
The StemLinc project, funded by the Marie Skłodowska-Curie Actions (MSCA) program, aimed to identify and characterize lncRNAs involved in the regulation of HSC stemness and their potential role in leukemia.
The project’s specific objectives were:
To generate a comprehensive catalog of hematopoietic lncRNAs through deep total RNA sequencing of primary mouse blood cells and bioinformatics analysis.
To infer the function of identified lncRNAs using computational approaches integrating genomic, epigenomic and transcriptomic data.
To validate the role of selected candidate lncRNAs through functional experiments and assess their relevance in leukemia.
By integrating advanced transcriptomics, functional genomics, and computational analyses, StemLinc significantly advances multiple scientific domains:
Hematopoietic system biology: Providing new insights into the transcriptional landscape of HSCs, progenitors, and differentiated blood cells.
ncRNA biology: Characterizing previously unexplored lncRNAs, including mono-exonic transcripts, broadening the understanding of non-coding RNA regulation in hematopoiesis.
Resource development for future research: Identifying candidate lncRNAs strongly associated with HSC identity and function, offering valuable insights for disease biomarker discovery and future large-scale studies.
The project followed a multidisciplinary approach, combining transcriptomics, computational biology, and functional genomics to achieve its objectives. The main activities and achievements included:
High-depth, strand-specific total RNA sequencing and transcriptome assembly of hematopoietic stem and progenitor cells (HSPCs) along with T cells and macrophages, leading to the identification of 1,779 novel lncRNAs.
Computational prediction of lncRNA functions, integrating intrinsic sequence features analysis, expression correlation, gene regulatory network analysis, transcription factor binding data, and chromatin interactions to infer functional roles of the identified lncRNAs.
Prioritization of a shortlist of lncRNAs strongly associated with HSC identity and function, based on their co-expression with key hematopoietic regulators and their presence in stem cell-enriched transcriptional networks.
Prioritization of a candidate lncRNA (XLOC_079903) for functional validation based on its association with the key hematopoietic regulator Kit and its predicted regulatory interactions.
Experimental validation using antisense oligonucleotide (ASO) knockdown, currently in progress, to determine the functional impact of the candidate lncRNA on HSC self-renewal and differentiation.
High-depth, strand-specific total RNA sequencing and transcriptome assembly of hematopoietic stem and progenitor cells (HSPCs) along with T cells and macrophages, leading to the identification of 1,779 novel lncRNAs.
Computational prediction of lncRNA functions, integrating intrinsic sequence features analysis, expression correlation, gene regulatory network analysis, transcription factor binding data, and chromatin interactions to infer functional roles of the identified lncRNAs.
Prioritization of a shortlist of lncRNAs strongly associated with HSC identity and function, based on their co-expression with key hematopoietic regulators and their presence in stem cell-enriched transcriptional networks.
Prioritization of a candidate lncRNA (XLOC_079903) for functional validation based on its association with the key hematopoietic regulator Kit and its predicted regulatory interactions.
Experimental validation using antisense oligonucleotide (ASO) knockdown, currently in progress, to determine the functional impact of the candidate lncRNA on HSC self-renewal and differentiation.
The StemLinc project has generated novel insights into the role of long non-coding RNAs (lncRNAs) in hematopoietic stem cell biology, expanding beyond previous knowledge in the field. The key breakthroughs include:
Expanding the known catalog of hematopoietic lncRNAs, including less-explored RNA types such as monoexonic lncRNAs. The dataset provides a new reference for hematopoiesis research, offering unique insights that go beyond existing publicly available resources.
Developing a novel integrative computational approach to infer lncRNA functions in hematopoiesis, providing a robust framework for future studies.
Uncovering a new lncRNA associated with the Kit gene, a crucial regulator of HSC function. This represents a strong candidate for controlling HSC homeostasis, potentially linking non-coding RNA regulation to essential stem cell pathways.
Providing a valuable dataset complementing existing resources. Our study is the first to perform RNA sequencing across various blood cells using total RNA, in contrast to the traditional approach of polyA-selected RNA-seq. This advancement not only enhances the identification of non-polyadenylated lncRNAs but also provides a novel approach to transcriptome analysis in hematopoiesis that could have broader implications for future studies.
While significant progress has been made, further research, validation, and translational efforts will be required to fully exploit these discoveries. Future steps will focus on:
Experimental validation of key lncRNA candidates in HSC function.
Assessing their role in human leukemia to explore clinical relevance.
Exploring IP protection and commercialization opportunities for potential applications in stem cell therapy and cancer treatment.
Further single-cell RNA sequencing (scRNA-seq) and epigenetic profiling could provide deeper insights into lncRNA function at the cellular level.
Expanding the known catalog of hematopoietic lncRNAs, including less-explored RNA types such as monoexonic lncRNAs. The dataset provides a new reference for hematopoiesis research, offering unique insights that go beyond existing publicly available resources.
Developing a novel integrative computational approach to infer lncRNA functions in hematopoiesis, providing a robust framework for future studies.
Uncovering a new lncRNA associated with the Kit gene, a crucial regulator of HSC function. This represents a strong candidate for controlling HSC homeostasis, potentially linking non-coding RNA regulation to essential stem cell pathways.
Providing a valuable dataset complementing existing resources. Our study is the first to perform RNA sequencing across various blood cells using total RNA, in contrast to the traditional approach of polyA-selected RNA-seq. This advancement not only enhances the identification of non-polyadenylated lncRNAs but also provides a novel approach to transcriptome analysis in hematopoiesis that could have broader implications for future studies.
While significant progress has been made, further research, validation, and translational efforts will be required to fully exploit these discoveries. Future steps will focus on:
Experimental validation of key lncRNA candidates in HSC function.
Assessing their role in human leukemia to explore clinical relevance.
Exploring IP protection and commercialization opportunities for potential applications in stem cell therapy and cancer treatment.
Further single-cell RNA sequencing (scRNA-seq) and epigenetic profiling could provide deeper insights into lncRNA function at the cellular level.