Biology subjects

Luc, S.

Publications and source records attributed to Luc, S..

6 recordsLinked to original sources

bioRxiv2023

Aging is associated with functional and molecular changes in distinct hematopoietic stem cell subsets

Age is a risk factor for hematologic malignancies. Attributes of the aging hematopoietic system include increased myelopoiesis, impaired adaptive immunity, and a functional decline of the hematopoietic stem cells (HSCs) that maintain hematopoiesis. Changes in the composition of diverse HSC subsets have been suggested to be responsible for age-related alterations, however, the underlying regulatory mechanisms are incompletely understood in the context of HSC heterogeneity. In this study, we investigated how distinct HSC subsets, separated by CD49b, functionally and molecularly change their behavior with age. We demonstrate that blood lineage differentiation progressively shifts to a higher myeloid cellular output in both lymphoid-biased and myeloid-biased HSC subsets during aging. In parallel, we show that HSCs selectively undergo age-dependent gene expression and gene regulatory molecular changes in a progressive manner, which is initiated already in the pre-adult stage. Overall, our studies suggest that aging intrinsically alters both cellular and molecular properties of HSCs. HighlightsO_LIWith age a gradual shift towards myeloid differentiation occurs in both myeloid-biased and lymphoid-biased enriched HSC subsets. C_LIO_LIAge-related molecular changes preferentially occur in HSCs. C_LIO_LIFunctionally distinct HSC subsets with high transcriptional similarity can be distinguished on the epigenetic level. C_LIO_LIHSC aging is associated with a progressive increase in chromatin accessibility. C_LI

cell biology↗

bioRxiv2023

An uncanonical CDK6 activity inhibits cilia function by suppressing axoneme polyglutamylation

Tubulin polyglutamylation is a post-translational modification that occurs primarily along the axoneme of cilia. Defective axoneme polyglutamylation impairs cilia function and has been correlated with ciliopathies, including Joubert Syndrome (JBTS). However, the precise mechanisms regulating proper axoneme polyglutamylation remain vague. Here, we show that Cyclin-Dependent Kinase 6 (CDK6), but not its paralog CDK4, localizes to cilia base and suppresses axoneme polyglutamylation by phosphorylating RAB11-interacting protein FIP5 at site S641, a critical regulator of cilia import of glutamylases. S641 phosphorylation disrupts the ciliary recruitment of FIP5 by impairing its association with RAB11, thereby reducing the ciliary import of glutamylases. Interestingly, significant upregulation of CDK6 and defective axoneme polyglutamylation were observed in Autosomal dominant polycystic kidney disease (ADPKD) cells. Encouragingly, the FDA-approved CDK4/6 inhibitor Abemaciclib can effectively restore cilia function in JBTS and ADPKD cells with defective glutamylation and suppresses renal cystogenesis in an ex vivo ADPKD model. In summary, our study elucidates regulatory mechanisms governing axoneme polyglutamylation and suggests developing CDK6-specific inhibitors could be a promising therapeutic strategy to enhance cilia function in ciliopathy patients.

cell biology↗

bioRxiv2023

Expression, not sequence, distinguishes miR-238 from its miR-239ab sister miRNAs in promoting longevity in Caenorhabditis elegans

MicroRNAs (miRNAs) regulate gene expression by base-pairing to target sequences in messenger RNAs (mRNAs) and recruiting factors that induce translational repression and mRNA decay. In animals, nucleotides 2-8 at the 5 end of the miRNA, called the seed region, are often necessary and sometimes sufficient for functional target interactions. MiRNAs that contain identical seed sequences are grouped into families where individual members have the potential to share targets and act redundantly. A rare exception seemed to be the miR-238/239ab family in Caenorhabditis elegans, as previous work indicated that loss of miR-238 reduced lifespan while deletion of the miR-239ab locus resulted in enhanced longevity and thermal stress resistance. Here, we re-examined these potentially opposing roles using new strains that individually disrupt each miRNA sister. We confirmed that loss of miR-238 is associated with a shortened lifespan but could detect no longevity or stress phenotypes in animals lacking miR-239a or miR-239b, individually or in combination. Additionally, dozens of genes were mis-regulated in miR-238 mutants but almost no gene expression changes were detected in either miR-239a or miR-239b mutants compared to wild type animals. We present evidence that the lack of redundancy between miR-238 and miR-239ab is independent of their sequence differences; miR-239a or miR-239b could substitute for the longevity role of miR-238 when expressed from the miR-238 locus. Altogether, these studies disqualify miR-239ab as negative regulators of aging and demonstrate that expression, not sequence, dictates the specific role of miR-238 in promoting longevity. Author SummaryMicroRNAs (miRNAs) are tiny non-coding RNAs that function in diverse biological pathways. To exert their regulatory influence, miRNAs bind to specific target RNAs through partial base-pairing. A critical aspect of this miRNA-target engagement is the seed sequence, nucleotides 2-8 of the miRNA. MiRNAs that share seed sequences are grouped into families and presumed to have similar functions. Yet, other factors, such as non-seed sequences in the miRNA and its expression level, can also contribute to target regulation and result in distinct roles for miRNAs within a family. To better understand how miRNA family members can have specific functions, we focused on miR-238 and its sisters, miR-239a and miR-239b, because these miRNAs had previously been reported to play opposing longevity roles in the nematode C. elegans. Using new genetic tools, we found that loss of miR-238 alone leads to the misregulation of many genes and a reduced lifespan. However, the lack of miR-239a, miR-239b, or both sisters had almost no effect on gene expression or longevity compared to wild type animals. Strikingly, though, miR-239a or miR-239b could substitute for the aging role of miR-238 when expressed from the miR-238 locus. Thus, expression, not sequence, is the predominant distinguishing feature of mir-238 that bestows upon it a role in aging not shared with the other family members.

genetics↗

bioRxiv2023

The combination of CD49b and CD229 reveals a subset of multipotent cells with short-term activity within the hematopoietic stem cell compartment

Hematopoiesis is maintained by hematopoietic stem cells (HSCs) that replenish all blood lineages throughout life. It is well-established that the HSC pool is functionally heterogeneous consisting of cells differing in longevity, self-renewal ability, cell proliferation, and lineage differentiation. Although HSCs can be identified through the Lineage-Sca-1+c-Kit+CD48-CD34- CD150+ immunophenotype, the cell surface marker combination does not permit absolute purification of functional HSCs with long-term reconstituting ability. Therefore, prospective isolation of long-term HSCs is crucial for mechanistic understanding of the biological functions of HSCs and for resolving functional heterogeneity within the HSC population. Here, we show that the combination of CD229 and CD49b cell surface markers within the phenotypic HSC compartment identifies a subset of multipotent progenitor cells with high proliferative activity and short-term reconstituting ability. Thus, the addition of CD229 and CD49b to conventional HSC markers permits prospective isolation of functional HSCs by distinguishing MPPs in the HSC compartment. SIGNIFICANCE STATEMENTBlood cells are generated by rare stem cells, which based on their defining features of self-renewal and multipotent abilities, are used in transplantation as curative treatment for blood diseases. Despite their clinical importance, mechanisms of stem cell properties are elusive given the inability to isolate pure blood stem cells for prospective analyses. In this study, we demonstrate that stem cells can be isolated by excluding multipotent progenitors, marked by CD229 and CD49b, from the stem cell compartment. Our study has important implications for the analysis of mechanisms regulating stem cell function and is relevant for clinical applications of stem cells.

cell biology↗

bioRxiv2022

T-RHEX-RNAseq - A tagmentation-based, rRNA blocked, random hexamer primed RNAseq method for generating stranded RNAseq libraries directly from very low numbers of lysed cells.

BackgroundRNA sequencing has become the mainstay for studies of gene expression. Still, analysis of rare cells with random hexamer priming - to allow analysis of a broader range of transcripts - remains challenging. ResultsWe here describe a tagmentation-based, rRNA blocked, random hexamer primed RNAseq approach (T-RHEX-RNAseq) for generating stranded RNAseq libraries from very low numbers of FACS sorted cells without RNA purification steps. ConclusionT-RHEX-RNAseq provides an easy-to-use, time efficient and automation compatible method for generating stranded RNAseq libraries from rare cells.

molecular biology↗

bioRxiv2022

FOXO dictates initiation of B cell development and myeloid restriction in common lymphoid progenitors

The development of B cells relies on an intricate network of transcription factors critical for developmental progression and lineage commitment. In the B cell developmental trajectory, a temporal switch from predominant Foxo3 to Foxo1 expression occurs at the CLP stage. Utilizing VAV-iCre mediated conditional deletion, we found that the loss of FOXO3 impaired B cell development from LMPP down to B cell precursors, while the loss of FOXO1 impaired B cell commitment and resulted in a complete developmental block at the CD25 negative proB cell stage. Strikingly, the combined loss of FOXO1 and FOXO3 resulted in the failure to restrict the myeloid potential of CLPs and the complete loss of the B cell lineage. This is underpinned by the failure to enforce the early B-lineage gene regulatory circuitry upon a predominantly pre-established open chromatin landscape. Altogether, this demonstrates that FOXO3 and FOXO1 cooperatively govern early lineage restriction and initiation of B-lineage commitment in CLPs. SUMMARYCommon lymphoid progenitors co-express the transcription factors FOXO1 and FOXO3. Removing FOXO1 and FOXO3 at this developmental stage results in regained myeloid potential, failed establishment of the early B cell gene regulatory program, and the complete loss of the B cell lineage.

immunology↗