Priscila Ferreira Slepicka

All Publications

• Functional Human iPSC-Derived Thymic Epithelial Progenitor Cells Reconstitute T Cell Development and Function in an In Vivo Model of Thymic Aplasia Slepicka, P., Hubka, K. M., Hanh Dan Nguyen, Mohammed, A., Wang, J., Gifford, C., Sebastiano, V., Weinacht, K. G. AMER SOC HEMATOLOGY. 2022: 7340-7341

  View details for DOI 10.1182/blood-2022-171133

  View details for Web of Science ID 000893230300154

• Sequential Stem Cell-Kidney Transplantation in Schimke Immuno-osseous Dysplasia. The New England journal of medicine Bertaina, A., Grimm, P. C., Weinberg, K., Parkman, R., Kristovich, K. M., Barbarito, G., Lippner, E., Dhamdhere, G., Ramachandran, V., Spatz, J. M., Fathallah-Shaykh, S., Atkinson, T. P., Al-Uzri, A., Aubert, G., van der Elst, K., Green, S. G., Agarwal, R., Slepicka, P. F., Shah, A. J., Roncarolo, M. G., Gallo, A., Concepcion, W., Lewis, D. B. 2022; 386 (24): 2295-2302

  Abstract

  Lifelong immunosuppression is required for allograft survival after kidney transplantation but may not ultimately prevent allograft loss resulting from chronic rejection. We developed an approach that attempts to abrogate immune rejection and the need for post-transplantation immunosuppression in three patients with Schimke immuno-osseous dysplasia who had both T-cell immunodeficiency and renal failure. Each patient received sequential transplants of alphabeta T-cell-depleted and CD19 B-cell-depleted haploidentical hematopoietic stem cells and a kidney from the same donor. Full donor hematopoietic chimerism and functional ex vivo T-cell tolerance was achieved, and the patients continued to have normal renal function without immunosuppression at 22 to 34 months after kidney transplantation. (Funded by the Kruzn for a Kure Foundation.).

  View details for DOI 10.1056/NEJMoa2117028

  View details for PubMedID 35704481

• Harnessing Mechanisms of Immune Tolerance to Improve Outcomes in Solid Organ Transplantation: A Review. Frontiers in immunology Slepicka, P. F., Yazdanifar, M., Bertaina, A. 2021; 12: 688460

  Abstract

  Survival after solid organ transplantation (SOT) is limited by chronic rejection as well as the need for lifelong immunosuppression and its associated toxicities. Several preclinical and clinical studies have tested methods designed to induce transplantation tolerance without lifelong immune suppression. The limited success of these strategies has led to the development of clinical protocols that combine SOT with other approaches, such as allogeneic hematopoietic stem cell transplantation (HSCT). HSCT prior to SOT facilitates engraftment of donor cells that can drive immune tolerance. Recent innovations in graft manipulation strategies and post-HSCT immune therapy provide further advances in promoting tolerance and improving clinical outcomes. In this review, we discuss conventional and unconventional immunological mechanisms underlying the development of immune tolerance in SOT recipients and how they can inform clinical advances. Specifically, we review the most recent mechanistic studies elucidating which immune regulatory cells dampen cytotoxic immune reactivity while fostering a tolerogenic environment. We further discuss how this understanding of regulatory cells can shape graft engineering and other therapeutic strategies to improve long-term outcomes for patients receiving HSCT and SOT.

  View details for DOI 10.3389/fimmu.2021.688460

  View details for PubMedID 34177941

• Pregnancy and Breast Cancer: Pathways to Understand Risk and Prevention TRENDS IN MOLECULAR MEDICINE Slepicka, P. F., Cyrill, S. L., dos Santos, C. O. 2019; 25 (10): 866–81

  Abstract

  Several studies have made strong efforts to understand how age and parity modulate the risk of breast cancer. A holistic understanding of the dynamic regulation of the morphological, cellular, and molecular milieu of the mammary gland offers insights into the drivers of breast cancer development as well as into potential prophylactic interventions, the latter being a longstanding ambition of the research and clinical community aspiring to eradicate the disease. In this review we discuss mechanisms that react to pregnancy signals, and we delineate the nuances of pregnancy-associated dynamism that contribute towards either breast cancer development or prevention. Further definition of the molecular basis of parity and breast cancer risk may allow the elaboration of tools to predict and survey those who are at risk of breast cancer development.

  View details for DOI 10.1016/j.molmed.2019.06.003

  View details for Web of Science ID 000490188500007

  View details for PubMedID 31383623

• BPTF Maintains Chromatin Accessibility and the Self-Renewal Capacity of Mammary Gland Stem Cells STEM CELL REPORTS Frey, W. D., Chaudhry, A., Slepicka, P. F., Ouellette, A. M., Kirberger, S. E., Pomerantz, W. K., Hannon, G. J., dos Santos, C. O. 2017; 9 (1): 23–31

  Abstract

  Chromatin remodeling is a key requirement for transcriptional control of cellular differentiation. However, the factors that alter chromatin architecture in mammary stem cells (MaSCs) are poorly understood. Here, we show that BPTF, the largest subunit of the NURF chromatin remodeling complex, is essential for MaSC self-renewal and differentiation of mammary epithelial cells (MECs). BPTF depletion arrests cells at a previously undefined stage of epithelial differentiation that is associated with an incapacity to achieve the luminal cell fate. Moreover, genome-wide analysis of DNA accessibility following genetic or chemical inhibition, suggests a role for BPTF in maintaining the open chromatin landscape at enhancers regions in MECs. Collectively, our study implicates BPTF in maintaining the unique epigenetic state of MaSCs.

  View details for DOI 10.1016/j.stemcr.2017.04.031

  View details for Web of Science ID 000405178400004

  View details for PubMedID 28579392

  View details for PubMedCentralID PMC5783326

• Functional Immune Tolerance Induced By Sequential Hematopoietic Stem Cell-Solid Organ Transplantation Bertaina, A., Barbarito, G., Ramachandran, V. V., Kristovich, K., Lippner, E., Fathallah-Shaykh, S., Al-Uzri, A., Shah, A. J., Aubert, G., Slepicka, P., Oppizzi, L., Agarwal, R., Roncarolo, M., Gallo, A., Concepcion, W., Weinberg, K. I., Parkman, R., Lewis, D. B., Grimm, P. C. AMER SOC HEMATOLOGY. 2021: 1818-+

  View details for DOI 10.1182/blood-2021-149920

  View details for Web of Science ID 000736398807056

• The molecular basis of mammary gland development and epithelial differentiation. Seminars in cell & developmental biology Slepicka, P. F., Somasundara, A. V., Dos Santos, C. O. 2020

  Abstract

  Our understanding of the molecular events underpinning the development of mammalian organ systems has been increasing rapidly in recent years. With the advent of new and improved next-generation sequencing methods, we are now able to dig deeper than ever before into the genomic and epigenomic events that play critical roles in determining the fates of stem and progenitor cells during the development of an embryo into an adult. In this review, we detail and discuss the genes and pathways that are involved in mammary gland development, from embryogenesis, through maturation into an adult gland, to the role of pregnancy signals in directing the terminal maturation of the mammary gland into a milk producing organ that can nurture the offspring. We also provide an overview of the latest research in the single-cell genomics of mammary gland development, which may help us to understand the lineage commitment of mammary stem cells (MaSCs) into luminal or basal epithelial cells that constitute the mammary gland. Finally, we summarize the use of 3D organoid cultures as a model system to study the molecular events during mammary gland development. Our increased investigation of the molecular requirements for normal mammary gland development will advance the discovery of targets to predict breast cancer risk and the development of new breast cancer therapies.

  View details for DOI 10.1016/j.semcdb.2020.09.014

  View details for PubMedID 33082117

• NEK10 interactome and depletion reveal new roles in mitochondria. Proteome science Peres de Oliveira, A., Basei, F. L., Slepicka, P. F., de Castro Ferezin, C., Melo-Hanchuk, T. D., de Souza, E. E., Lima, T. I., Dos Santos, V. T., Mendes, D., Silveira, L. R., Menck, C. F., Kobarg, J. 2020; 18: 4

  Abstract

  Members of the family of NEK protein kinases (NIMA-related kinases) were described to have crucial roles in regulating different aspects of the cell cycle. NEK10 was reported to take part in the maintenance of the G2/M checkpoint after exposure to ultraviolet light. NEK1, NEK5, NEK2 and NEK4 proteins on the other hand have been linked to mitochondrial functions.HEK293T cells were transfected with FLAG empty vector or FLAG-NEK10 and treated or not with Zeocin. For proteomic analysis, proteins co-precipitated with the FLAG constructs were digested by trypsin, and then analyzed via LC-MS/MS. Proteomic data retrieved were next submitted to Integrated Interactome System analysis and differentially expressed proteins were attributed to Gene Ontology biological processes and assembled in protein networks by Cytoscape. For functional, cellular and molecular analyses two stable Nek10 silenced HeLa cell clones were established.Here, we discovered the following possible new NEK10 protein interactors, related to mitochondrial functions: SIRT3, ATAD3A, ATAD3B, and OAT. After zeocin treatment, the spectrum of mitochondrial interactors increased by the proteins: FKBP4, TXN, PFDN2, ATAD3B, MRPL12, ATP5J, DUT, YWHAE, CS, SIRT3, HSPA9, PDHB, GLUD1, DDX3X, and APEX1. We confirmed the interaction of NEK10 and GLUD1 by proximity ligation assay and confocal microscopy. Furthermore, we demonstrated that NEK10-depleted cells showed more fragmented mitochondria compared to the control cells. The knock down of NEK10 resulted further in changes in mitochondrial reactive oxygen species (ROS) levels, decreased citrate synthase activity, and culminated in inhibition of mitochondrial respiration, affecting particularly ATP-linked oxygen consumption rate and spare capacity. NEK10 depletion also decreased the ratio of mtDNA amplification, possibly due to DNA damage. However, the total mtDNA content increased, suggesting that NEK10 may be involved in the control of mtDNA content.Taken together these data place NEK10 as a novel regulatory player in mitochondrial homeostasis and energy metabolism.

  View details for DOI 10.1186/s12953-020-00160-w

  View details for PubMedID 32368190

  View details for PubMedCentralID PMC7189645

• NEK5 interacts with topoisomerase II beta and is involved in the DNA damage response induced by etoposide JOURNAL OF CELLULAR BIOCHEMISTRY Melo-Hanchuk, T., Slepicka, P., Pelegrini, A., Martins Menck, C., Kobarg, J. 2019; 120 (10): 16853–66

  Abstract

  Cells are daily submitted to high levels of DNA lesions that trigger complex pathways and cellular responses by cell cycle arrest, apoptosis, alterations in transcriptional response, and the onset of DNA repair. Members of the NIMA-related kinase (NEK) family have been related to DNA damage response and repair and the first insight about NEK5 in this context is related to its role in centrosome separation resulting in defects in chromosome integrity. Here we investigate the potential correlation between NEK5 and the DNA damage repair index. The effect of NEK5 in double-strand breaks caused by etoposide was accessed by alkaline comet assay and revealed that NEK5-silenced cells are more sensitive to etoposide treatment. Topoisomerase IIβ (TOPIIβ) is a target of etoposide that leads to the production of DNA breaks. We demonstrate that NEK5 interacts with TOPIIβ, and the dynamics of this interaction is evaluated by proximity ligation assay. The complex NEK5/TOPIIβ is formed immediately after etoposide treatment. Taken together, the results of our study reveal that NEK5 depletion increases DNA damage and impairs proper DNA damage response, pointing out NEK5 as a potential kinase contributor to genomic stability.

  View details for DOI 10.1002/jcb.28943

  View details for Web of Science ID 000483551100050

  View details for PubMedID 31090963

• NEK1 kinase domain structure and its dynamic protein interactome after exposure to Cisplatin SCIENTIFIC REPORTS Melo-Hanchuk, T. D., Slepicka, P., Meirelles, G., Basei, F., Lovato, D., Granato, D., Pauletti, B., Domingues, R., Paes Leme, A., Pelegrini, A., Lenz, G., Knapp, S., Elkins, J. M., Kobarg, J. 2017; 7: 5445

  Abstract

  NEK family kinases are serine/threonine kinases that have been functionally implicated in the regulation of the disjunction of the centrosome, the assembly of the mitotic spindle, the function of the primary cilium and the DNA damage response. NEK1 shows pleiotropic functions and has been found to be mutated in cancer cells, ciliopathies such as the polycystic kidney disease, as well as in the genetic diseases short-rib thoracic dysplasia, Mohr-syndrome and amyotrophic lateral sclerosis. NEK1 is essential for the ionizing radiation DNA damage response and priming of the ATR kinase and of Rad54 through phosphorylation. Here we report on the structure of the kinase domain of human NEK1 in its apo- and ATP-mimetic inhibitor bound forms. The inhibitor bound structure may allow the design of NEK specific chemo-sensitizing agents to act in conjunction with chemo- or radiation therapy of cancer cells. Furthermore, we characterized the dynamic protein interactome of NEK1 after DNA damage challenge with cisplatin. Our data suggest that NEK1 and its interaction partners trigger the DNA damage pathways responsible for correcting DNA crosslinks.

  View details for DOI 10.1038/s41598-017-05325-w

  View details for Web of Science ID 000405464200082

  View details for PubMedID 28710492

  View details for PubMedCentralID PMC5511132

• Nek5 interacts with mitochondrial proteins and interferes negatively in mitochondrial mediated cell death and respiration CELLULAR SIGNALLING Melo Hanchuk, T. D., Papa, P., La Guardia, P. G., Vercesi, A. E., Kobarg, J. 2015; 27 (6): 1168–77

  Abstract

  Mitochondria are involved in energy supply, signaling, cell death and cellular differentiation and have been implicated in several human diseases. Neks (NIMA-related kinases) represent a family of mammal protein kinases that play essential roles in cell-cycle progression, but other functions have recently been related. A yeast two-hybrid (Y2H) screen was performed to identify and characterize Nek5 interaction partners and the mitochondrial proteins Cox11, MTX-2 and BCLAF1 were retrieved. Apoptosis assay showed protective effects of stable hNek5 expression from Hek293-T's cell death after thapsigargin treatment (2 μM). Nek5 silenced cells as well as cells expressing a "kinase dead" version of Nek5, displayed an increase in ROS formation after 4 h of thapsigargin treatment. Mitochondrial respiratory chain activity was found decreased upon stable hNek5expression. Cells silenced for hNek5 on the other hand presented 1.7 fold increased basal rates of respiration, especially at the electrons transfer steps from TMPD to cytochrome c and at the complex II. In conclusion, our data suggest for the first time mitochondrial localization and functions for Nek5 and its participation in cell death and cell respiration regulation. Stable expression of hNek5 in Hek293T cells resulted in enhanced cell viability, decreased cell death and drug resistance, while depletion of hNek5by shRNA overcame cancer cell drug resistance and induced apoptosis in vitro. Stable expression of hNek5 also inhibits thapsigargin promoted apoptosis and the respiratory chain complex IV in HEK293T cells.

  View details for DOI 10.1016/j.cellsig.2015.02.021

  View details for Web of Science ID 000353096700013

  View details for PubMedID 25725288

• "Stop Ne(c)king around": How interactomics contributes to functionally characterize Nek family kinases. World journal of biological chemistry Meirelles, G. V., Perez, A. M., de Souza, E. E., Basei, F. L., Papa, P. F., Melo Hanchuk, T. D., Cardoso, V. B., Kobarg, J. 2014; 5 (2): 141-60

  Abstract

  Aside from Polo and Aurora, a third but less studied kinase family involved in mitosis regulation is the never in mitosis-gene A (NIMA)-related kinases (Neks). The founding member of this family is the sole member NIMA of Aspergillus nidulans, which is crucial for the initiation of mitosis in that organism. All 11 human Neks have been functionally assigned to one of the three core functions established for this family in mammals: (1) centrioles/mitosis; (2) primary ciliary function/ciliopathies; and (3) DNA damage response (DDR). Recent findings, especially on Nek 1 and 8, showed however, that several Neks participate in parallel in at least two of these contexts: primary ciliary function and DDR. In the core section of this in-depth review, we report the current detailed functional knowledge on each of the 11 Neks. In the discussion, we return to the cross-connections among Neks and point out how our and other groups' functional and interactomics studies revealed that most Neks interact with protein partners associated with two if not all three of the functional contexts. We then raise the hypothesis that Neks may be the connecting regulatory elements that allow the cell to fine tune and synchronize the cellular events associated with these three core functions. The new and exciting findings on the Nek family open new perspectives and should allow the Neks to finally claim the attention they deserve in the field of kinases and cell cycle biology.

  View details for DOI 10.4331/wjbc.v5.i2.141

  View details for PubMedID 24921005

  View details for PubMedCentralID PMC4050109

• Solution structure of the human signaling protein RACK1 BMC STRUCTURAL BIOLOGY Goncalves, K. A., Borges, J. C., Silva, J. C., Papa, P. F., Bressan, G. C., Torriani, I. L., Kobarg, J. 2010; 10: 15

  Abstract

  The adaptor protein RACK1 (receptor of activated kinase 1) was originally identified as an anchoring protein for protein kinase C. RACK1 is a 36 kDa protein, and is composed of seven WD repeats which mediate its protein-protein interactions. RACK1 is ubiquitously expressed and has been implicated in diverse cellular processes involving: protein translation regulation, neuropathological processes, cellular stress, and tissue development.In this study we performed a biophysical analysis of human RACK1 with the aim of obtaining low resolution structural information. Small angle X-ray scattering (SAXS) experiments demonstrated that human RACK1 is globular and monomeric in solution and its low resolution structure is strikingly similar to that of an homology model previously calculated by us and to the crystallographic structure of RACK1 isoform A from Arabidopsis thaliana. Both sedimentation velocity and sedimentation equilibrium analytical ultracentrifugation techniques showed that RACK1 is predominantly a monomer of around 37 kDa in solution, but also presents small amounts of oligomeric species. Moreover, hydrodynamic data suggested that RACK1 has a slightly asymmetric shape. The interaction of RACK1 and Ki-1/57 was tested by sedimentation equilibrium. The results suggested that the association between RACK1 and Ki-1/57(122-413) follows a stoichiometry of 1:1. The binding constant (KB) observed for RACK1-Ki-1/57(122-413) interaction was of around (1.5 +/- 0.2) x 10(6) M(-1) and resulted in a dissociation constant (KD) of (0.7 +/- 0.1) x 10(-6) M. Moreover, the fluorescence data also suggests that the interaction may occur in a cooperative fashion.Our SAXS and analytical ultracentrifugation experiments indicated that RACK1 is predominantly a monomer in solution. RACK1 and Ki-1/57(122-413) interact strongly under the tested conditions.

  View details for DOI 10.1186/1472-6807-10-15

  View details for Web of Science ID 000279917600002

  View details for PubMedID 20529362

  View details for PubMedCentralID PMC2896345