Ewa Bielczyk Maczynska

All Publications

• Single-cell transcriptome dataset of human and mouse in vitro adipogenesis models. Scientific data Li, J., Jin, C., Gustafsson, S., Rao, A., Wabitsch, M., Park, C. Y., Quertermous, T., Knowles, J. W., Bielczyk-Maczynska, E. 2023; 10 (1): 387

  Abstract

  Adipogenesis is a process in which fat-specific progenitor cells (preadipocytes) differentiate into adipocytes that carry out the key metabolic functions of the adipose tissue, including glucose uptake, energy storage, and adipokine secretion. Several cell lines are routinely used to study the molecular regulation of adipogenesis, in particular the immortalized mouse 3T3-L1 line and the primary human Simpson-Golabi-Behmel syndrome (SGBS) line. However, the cell-to-cell variability of transcriptional changes prior to and during adipogenesis in these models is not well understood. Here, we present a single-cell RNA-Sequencing (scRNA-Seq) dataset collected before and during adipogenic differentiation of 3T3-L1 and SGBS cells. To minimize the effects of experimental variation, we mixed 3T3-L1 and SGBS cells and used computational analysis to demultiplex transcriptomes of mouse and human cells. In both models, adipogenesis results in the appearance of three cell clusters, corresponding to preadipocytes, early and mature adipocytes. These data provide a groundwork for comparative studies on these widely used in vitro models of human and mouse adipogenesis, and on cell-to-cell variability during this process.

  View details for DOI 10.1038/s41597-023-02293-x

  View details for PubMedID 37328521

  View details for PubMedCentralID 2597101

• Single-cell transcriptome dataset of human and mouse in vitro adipogenesis models. bioRxiv : the preprint server for biology Li, J., Jin, C., Gustafsson, S., Rao, A., Wabitsch, M., Park, C. Y., Quertermous, T., Bielczyk-Maczynska, E., Knowles, J. W. 2023

  Abstract

  Adipogenesis is a process in which fat-specific progenitor cells (preadipocytes) differentiate into adipocytes that carry out the key metabolic functions of the adipose tissue, including glucose uptake, energy storage, and adipokine secretion. Several cell lines are routinely used to study the molecular regulation of adipogenesis, in particular the immortalized mouse 3T3-L1 line and the primary human Simpson-Golabi-Behmel syndrome (SGBS) line. However, the cell-to-cell variability of transcriptional changes prior to and during adipogenesis in these models is not well understood. Here, we present a single-cell RNA-Sequencing (scRNA-Seq) dataset collected before and during adipogenic differentiation of 3T3-L1 and SGBS cells. To minimize the effects of experimental variation, we mixed 3T3-L1 and SGBS cells and used computational analysis to demultiplex transcriptomes of mouse and human cells. In both models, adipogenesis results in the appearance of three cell clusters, corresponding to preadipocytes, early and mature adipocytes. These data provide a groundwork for comparative studies on human and mouse adipogenesis, as well as on cell-to-cell variability in gene expression during this process.

  View details for DOI 10.1101/2023.03.27.534456

  View details for PubMedID 37034809

  View details for PubMedCentralID PMC10081256

• G protein-coupled receptor 151 regulates glucose metabolism and hepatic gluconeogenesis Nature Communications Bielczyk-Maczynska, E., Zhao, M., Zushin, P. H., Schnurr, T. M., Kim, H., Li, J., Sangwung, P., Nallagatla, P., Park, C., Cornn, C., Stahl, A., Svensson, K. J., Knowles, J. W. 2022

  View details for DOI 10.1038/s41467-022-35069-9

• TGF-beta is insufficient to induce adipocyte state loss without concurrent PPARgamma downregulation. Scientific reports Taylor, B., Shah, A., Bielczyk-Maczynska, E. 2020; 10 (1): 14084

  Abstract

  Cell plasticity, the ability of differentiated cells to convert into other cell types, underlies the pathogenesis of many diseases including the transdifferentiation of adipocytes (fat cells) into myofibroblasts in the pathogenesis of dermal fibrosis. Loss of adipocyte identity is an early step in different types of adipocyte plasticity. In this study, we determine the dynamics of adipocyte state loss in response to the profibrotic cytokine TGF-beta. We use two complementary approaches, lineage tracing and live fluorescent microscopy, which both allow for robust quantitative tracking of adipocyte identity loss at the single-cell level. We find that the intracellular TGF-beta signaling in adipocytes is inhibited by the transcriptional factor PPARgamma, specifically by its ubiquitously expressed isoform PPARgamma1. However, TGF-beta can lead to adipocyte state loss when it is present simultaneously with another stimulus. Our findings establish that an integration of stimuli occurring in a specific order is pivotal for adipocyte state loss which underlies adipocyte plasticity. Our results also suggest the possibility of a more general switch-like mechanism between adipogenic and profibrotic molecular states.

  View details for DOI 10.1038/s41598-020-71100-z

  View details for PubMedID 32826933

• White Adipocyte Plasticity in Physiology and Disease. Cells Bielczyk-Maczynska, E. n. 2019; 8 (12)

  Abstract

  Cellular plasticity is a transformation of a terminally differentiated cell into another cell type, which has been long known to occur in disease and regeneration. However, white adipocytes (fat cells) have only recently been observed to undergo different types of cellular plasticity. Adipocyte transdifferentiation into myofibroblasts and cancer-associated fibroblasts occurs in fibrosis and cancer, respectively. On the other hand, reversible adipocyte dedifferentiation into adipocyte progenitor cells (preadipocytes) has been demonstrated in mammary gland and in dermal adipose tissue. Here we discuss the research on adipocyte plasticity, including the experimental approaches that allowed to detect and study it, the current state of the knowledge, major research questions which remain to be addressed, and the advances required to stimulate adipocyte plasticity research. In the future, the knowledge of the molecular mechanisms of adipocyte plasticity can be utilized both to prevent adipocyte plasticity in disease and to stimulate it for use in regenerative medicine.

  View details for DOI 10.3390/cells8121507

  View details for PubMedID 31775295

• The Ribosome Biogenesis Protein Nol9 Is Essential for Definitive Hematopoiesis and Pancreas Morphogenesis in Zebrafish PLOS GENETICS Bielczyk-Maczynska, E., Hung, L. L., Ferreira, L., Fleischmann, T., Weis, F., Fernandez-Pevida, A., Harvey, S. A., Wali, N., Warren, A. J., Barroso, I., Stemple, D. L., Cvejic, A. 2015; 11 (12)

  Abstract

  Ribosome biogenesis is a ubiquitous and essential process in cells. Defects in ribosome biogenesis and function result in a group of human disorders, collectively known as ribosomopathies. In this study, we describe a zebrafish mutant with a loss-of-function mutation in nol9, a gene that encodes a non-ribosomal protein involved in rRNA processing. nol9sa1022/sa1022 mutants have a defect in 28S rRNA processing. The nol9sa1022/sa1022 larvae display hypoplastic pancreas, liver and intestine and have decreased numbers of hematopoietic stem and progenitor cells (HSPCs), as well as definitive erythrocytes and lymphocytes. In addition, ultrastructural analysis revealed signs of pathological processes occurring in endothelial cells of the caudal vein, emphasizing the complexity of the phenotype observed in nol9sa1022/sa1022 larvae. We further show that both the pancreatic and hematopoietic deficiencies in nol9sa1022/sa1022 embryos were due to impaired cell proliferation of respective progenitor cells. Interestingly, genetic loss of Tp53 rescued the HSPCs but not the pancreatic defects. In contrast, activation of mRNA translation via the mTOR pathway by L-Leucine treatment did not revert the erythroid or pancreatic defects. Together, we present the nol9sa1022/sa1022 mutant, a novel zebrafish ribosomopathy model, which recapitulates key human disease characteristics. The use of this genetically tractable model will enhance our understanding of the tissue-specific mechanisms following impaired ribosome biogenesis in the context of an intact vertebrate.

  View details for DOI 10.1371/journal.pgen.1005677

  View details for Web of Science ID 000368518400017

  View details for PubMedID 26624285

• A Loss of Function Screen of Identified Genome-Wide Association Study Loci Reveals New Genes Controlling Hematopoiesis PLOS GENETICS Bielczyk-Maczynska, E., Serbanovic-Canic, J., Ferreira, L., Soranzo, N., Stemple, D. L., Ouwehand, W. H., Cvejic, A. 2014; 10 (7)

  Abstract

  The formation of mature cells by blood stem cells is very well understood at the cellular level and we know many of the key transcription factors that control fate decisions. However, many upstream signalling and downstream effector processes are only partially understood. Genome wide association studies (GWAS) have been particularly useful in providing new directions to dissect these pathways. A GWAS meta-analysis identified 68 genetic loci controlling platelet size and number. Only a quarter of those genes, however, are known regulators of hematopoiesis. To determine function of the remaining genes we performed a medium-throughput genetic screen in zebrafish using antisense morpholino oligonucleotides (MOs) to knock down protein expression, followed by histological analysis of selected genes using a wide panel of different hematopoietic markers. The information generated by the initial knockdown was used to profile phenotypes and to position candidate genes hierarchically in hematopoiesis. Further analysis of brd3a revealed its essential role in differentiation but not maintenance and survival of thrombocytes. Using the from-GWAS-to-function strategy we have not only identified a series of genes that represent novel regulators of thrombopoiesis and hematopoiesis, but this work also represents, to our knowledge, the first example of a functional genetic screening strategy that is a critical step toward obtaining biologically relevant functional data from GWA study for blood cell traits.

  View details for DOI 10.1371/journal.pgen.1004450

  View details for Web of Science ID 000339902600011

  View details for PubMedID 25010335

• A single-cell CRISPRi platform for characterizing candidate genes relevant to metabolic disorders in human adipocytes. American journal of physiology. Cell physiology Bielczyk-Maczynska, E., Sharma, D., Blencowe, M., Saliba Gustafsson, P., Gloudemans, M. J., Yang, X., Carcamo-Orive, I., Wabitsch, M., Svensson, K. J., Park, C. Y., Quertermous, T., Knowles, J. W., Li, J. 2023

  Abstract

  CROP-Seq combines gene silencing using CRISPR interference with single-cell RNA sequencing. Here, we applied CROP-Seq to study adipogenesis and adipocyte biology. Human preadipocyte SGBS cell line expressing KRAB-dCas9 was transduced with a sgRNA library. Following selection, individual cells were captured using microfluidics at different timepoints during adipogenesis. Bioinformatic analysis of transcriptomic data was used to determine the knock-down effects, the dysregulated pathways, and to predict cellular phenotypes. Single-cell transcriptomes recapitulated adipogenesis states. For all targets, over 400 differentially expressed genes were identified at least at one timepoint. As a validation of our approach, the knock-down of PPARG and CEBPB (which encode key proadipogenic transcription factors) resulted in the inhibition of adipogenesis. Gene set enrichment analysis generated hypotheses regarding the molecular function of novel genes. MAFF knock-down led to downregulation of transcriptional response to proinflammatory cytokine TNF-α in preadipocytes and to decreased CXCL-16 and IL-6 secretion. TIPARP knock-down resulted in increased expression of adipogenesis markers. In summary, this powerful, hypothesis-free tool can identify novel regulators of adipogenesis, preadipocyte and adipocyte function associated with metabolic disease.

  View details for DOI 10.1152/ajpcell.00148.2023

  View details for PubMedID 37486064

• Flattening of circadian glucocorticoid oscillations drives acute hyperinsulinemia and adipocyte hypertrophy. Cell reports Tholen, S., Patel, R., Agas, A., Kovary, K. M., Rabiee, A., Nicholls, H. T., Bielczyk-Maczyńska, E., Yang, W., Kraemer, F. B., Teruel, M. N. 2022; 39 (13): 111018

  Abstract

  Disruption of circadian glucocorticoid oscillations in Cushing's disease and chronic stress results in obesity and adipocyte hypertrophy, which is believed to be a main source of the harmful effects of obesity. Here, we recapitulate stress due to jet lag or work-life imbalances by flattening glucocorticoid oscillations in mice. Within 3 days, mice achieve a metabolic state with persistently high insulin, but surprisingly low glucose and fatty acids in the bloodstream, that precedes a more than 2-fold increase in brown and white adipose tissue mass within 3 weeks. Transcriptomic and Cd36-knockout mouse analyses show that hyperinsulinemia-mediated de novo fatty acid synthesis and Cd36-mediated fatty acid uptake drive fat mass increases. Intriguingly, this mechanism by which glucocorticoid flattening causes acute hyperinsulinemia and adipocyte hypertrophy is unexpectedly beneficial in preventing high levels of circulating fatty acids and glucose for weeks, thus serving as a protective response to preserve metabolic health during chronic stress.

  View details for DOI 10.1016/j.celrep.2022.111018

  View details for PubMedID 35767959

• Phosphoproteomic mapping reveals distinct signaling actions and activation of muscle protein synthesis by Isthmin-1 eLife Zhao, M., Banhos Danneskiold-Samsøe, N., Ulicna, L., Nguyen, Q., Voilquin, L., Lee, D. E., White, J. P., Jiang, Z., Cuthbert, N., Paramasivam, S., Bielczyk-Maczynska, E., van Rechem, C., Svensson, K. J. 2022

  View details for DOI 10.7554/eLife.80014

• Loss of the homologous recombination gene rad51 leads to Fanconi anemia-like symptoms in zebrafish. Proceedings of the National Academy of Sciences of the United States of America Botthof, J. G., Bielczyk-Maczyńska, E. n., Ferreira, L. n., Cvejic, A. n. 2017; 114 (22): E4452–E4461

  Abstract

  RAD51 is an indispensable homologous recombination protein, necessary for strand invasion and crossing over. It has recently been designated as a Fanconi anemia (FA) gene, following the discovery of two patients carrying dominant-negative mutations. FA is a hereditary DNA-repair disorder characterized by various congenital abnormalities, progressive bone marrow failure, and cancer predisposition. In this report, we describe a viable vertebrate model of RAD51 loss. Zebrafish rad51 loss-of-function mutants developed key features of FA, including hypocellular kidney marrow, sensitivity to cross-linking agents, and decreased size. We show that some of these symptoms stem from both decreased proliferation and increased apoptosis of embryonic hematopoietic stem and progenitor cells. Comutation of p53 was able to rescue the hematopoietic defects seen in the single mutants, but led to tumor development. We further demonstrate that prolonged inflammatory stress can exacerbate the hematological impairment, leading to an additional decrease in kidney marrow cell numbers. These findings strengthen the assignment of RAD51 as a Fanconi gene and provide more evidence for the notion that aberrant p53 signaling during embryogenesis leads to the hematological defects seen later in life in FA. Further research on this zebrafish FA model will lead to a deeper understanding of the molecular basis of bone marrow failure in FA and the cellular role of RAD51.

  View details for DOI 10.1073/pnas.1620631114

  View details for PubMedID 28512217

  View details for PubMedCentralID PMC5465903

• Transcriptional diversity during lineage commitment of human blood progenitors SCIENCE Chen, L., Kostadima, M., Martens, J. H., Canu, G., Garcia, S. P., Turro, E., Downes, K., Macaulay, I. C., Bielczyk-Maczynska, E., Coe, S., Farrow, S., Poudel, P., Burden, F., Jansen, S. B., Astle, W. J., Attwood, A., Bariana, T., de Bono, B., Breschi, A., Chambers, J. C., Choudry, F. A., Clarke, L., Coupland, P., van der Ent, M., Erber, W. N., Jansen, J. H., Favier, R., Fenech, M. E., Foad, N., Freson, K., Van Geet, C., Gomez, K., Guigo, R., Hampshire, D., Kelly, A. M., Kerstens, H. H., Kooner, J. S., Laffan, M., Lentaigne, C., Labalette, C., Martin, T., Meacham, S., Mumford, A., Nuernberg, S., Palumbo, E., van der Reijden, B. A., Richardson, D., Sammut, S. J., Slodkowicz, G., Tamuri, A. U., Vasquez, L., Voss, K., Watt, S., Westbury, S., Flicek, P., Loos, R., Goldman, N., Bertone, P., Read, R. J., Richardson, S., Cvejic, A., Soranzo, N., Ouwehand, W. H., Stunnenberg, H. G., Frontini, M., Rendon, A. 2014; 345 (6204): 1580-?

  Abstract

  Blood cells derive from hematopoietic stem cells through stepwise fating events. To characterize gene expression programs driving lineage choice, we sequenced RNA from eight primary human hematopoietic progenitor populations representing the major myeloid commitment stages and the main lymphoid stage. We identified extensive cell type-specific expression changes: 6711 genes and 10,724 transcripts, enriched in non-protein-coding elements at early stages of differentiation. In addition, we found 7881 novel splice junctions and 2301 differentially used alternative splicing events, enriched in genes involved in regulatory processes. We demonstrated experimentally cell-specific isoform usage, identifying nuclear factor I/B (NFIB) as a regulator of megakaryocyte maturation-the platelet precursor. Our data highlight the complexity of fating events in closely related progenitor populations, the understanding of which is essential for the advancement of transplantation and regenerative medicine.

  View details for DOI 10.1126/science.1251033

  View details for Web of Science ID 000342164500035

  View details for PubMedID 25258084

• SMIM1 underlies the Vel blood group and influences red blood cell traits NATURE GENETICS Cvejic, A., Haer-Wigman, L., Stephens, J. C., Kostadima, M., Smethurst, P. A., Frontini, M., Van den Akker, E., Bertone, P., Bielczyk-Maczynska, E., Farrow, S., Fehrmann, R. S., Gray, A., de Haas, M., Haver, V. G., Jordan, G., Karjalainen, J., Kerstens, H. H., Kiddle, G., Lloyd-Jones, H., Needs, M., Poole, J., Soussan, A. A., Rendon, A., Rieneck, K., Sambrook, J. G., Schepers, H., Sillje, H. H., Sipos, B., Swinkels, D., Tamuri, A. U., Verweij, N., Watkins, N. A., Westra, H., Stemple, D., Franke, L., Soranzo, N., Stunnenberg, H. G., Goldman, N., van der Harst, P., van der Schoot, C. E., Ouwehand, W. H., Albers, C. A. 2013; 45 (5): 542-U115

  Abstract

  The blood group Vel was discovered 60 years ago, but the underlying gene is unknown. Individuals negative for the Vel antigen are rare and are required for the safe transfusion of patients with antibodies to Vel. To identify the responsible gene, we sequenced the exomes of five individuals negative for the Vel antigen and found that four were homozygous and one was heterozygous for a low-frequency 17-nucleotide frameshift deletion in the gene encoding the 78-amino-acid transmembrane protein SMIM1. A follow-up study showing that 59 of 64 Vel-negative individuals were homozygous for the same deletion and expression of the Vel antigen on SMIM1-transfected cells confirm SMIM1 as the gene underlying the Vel blood group. An expression quantitative trait locus (eQTL), the common SNP rs1175550 contributes to variable expression of the Vel antigen (P = 0.003) and influences the mean hemoglobin concentration of red blood cells (RBCs; P = 8.6 × 10(-15)). In vivo, zebrafish with smim1 knockdown showed a mild reduction in the number of RBCs, identifying SMIM1 as a new regulator of RBC formation. Our findings are of immediate relevance, as the homozygous presence of the deletion allows the unequivocal identification of Vel-negative blood donors.

  View details for DOI 10.1038/ng.2603

  View details for Web of Science ID 000318158200018

  View details for PubMedID 23563608