Felicia Reinitz

All Publications

• Basic Science and Pathogenesis. Alzheimer's & dementia : the journal of the Alzheimer's Association Ho, W. H., Djomehri, S. I., Alford, V. M., Chen, E. Y., Reinitz, F., Gottlieb, S., Igboabuchi, F. I., Chung, G. Y., Fernandez, D., Montabana, E., Singal, B., Qi, Z., Kuo, A. H., Qian, D., Clarke, M. F. 2024; 20 Suppl 1: e093300

  Abstract

  Hallmark pathologies of Alzheimer's Disease (AD) include the accumulation of both extracellular amyloid and intracellular tau proteins. While a significant body of knowledge exists surrounding the role of the protein aggregates in the context of AD, research supporting these as targets for therapeutic development have yielded inconsistent findings. One significant barrier is the inability to restore cognitive function despite the successful clearance of these proteins. In our recently published paper, we discuss a novel target for AD, USP16, which has been shown to promote ubiquitination of histones allowing for transcription of CDKN2A, a cell cycle regulator. We demonstrated that knockdown of USP16 improved neurosphere initiating capacity in mouse neural stem cells. Moreover, reduction of USP16 expression via heterozygosity resulted in improved cognition in 12-month-old mice. From these findings, our lab aims to identify the full-length protein structure of USP16 using cryogenic electron microscopy (cryo-EM) and develop a small molecule drug to inhibit its activity for end-use as a viable AD therapeutic option.Truncated regions corresponding to the active and zinc-finger (ZnF) domains of USP16 were expressed in both HEK293T and SF9 cells. Whole cell lysates were purified using affinity chromatography and identity was verified using Western blot. The purified protein products were submitted for cryo-EM to determine their respective structures. Focusing on the ZnF domain, we utilized both a previously published proton NMR-derived structure as well as AlphaFold-predicted USP16 protein structure to serve as model scaffolding onto which we could perform in silico small molecule drug screening. Libraries were obtained from Schrodinger, MilliporeSigma, Emanine, MolPort, Thermo Fisher, MedChemExpress, Life Chemicals, ChemDiv, and Mcule.Chromatogram from affinity chromatography and Western blot imaging together indicate successful isolation of truncated USP16 domains expressed in HEK293T and SF9 cells. Preliminary findings from cryo-EM on the WT SF9 construct recapitulates the major domains predicted by AlphaFold. Over 9.5 million small molecule hit interactions against the ZnF domain have been identified.Cryo-EM was successful in generating a course model of USP16. In silico protein modeling and drug screening was able to identify potential ligands against the ZnF domain of USP16.

  View details for DOI 10.1002/alz.093300

  View details for PubMedID 39750542

• Inhibiting USP16 rescues stem cell aging and memory in an Alzheimer's model. eLife Reinitz, F., Chen, E. Y., Nicolis di Robilant, B., Chuluun, B., Antony, J., Jones, R. C., Gubbi, N., Lee, K., Ho, W. H., Kolluru, S. S., Qian, D., Adorno, M., Piltti, K., Anderson, A., Monje, M., Heller, H. C., Quake, S. R., Clarke, M. F. 2022; 11

  Abstract

  Alzheimer's disease (AD) is a progressive neurodegenerative disease observed with aging that represents the most common form of dementia. To date, therapies targeting end-stage disease plaques, tangles, or inflammation have limited efficacy. Therefore, we set out to identify a potential earlier targetable phenotype. Utilizing a mouse model of AD and human fetal cells harboring mutant amyloid precursor protein, we show cell intrinsic neural precursor cell (NPC) dysfunction precedes widespread inflammation and amyloid plaque pathology, making it the earliest defect in the evolution of the disease. We demonstrate that reversing impaired NPC self-renewal via genetic reduction of USP16, a histone modifier and critical physiological antagonist of the Polycomb Repressor Complex 1, can prevent downstream cognitive defects and decrease astrogliosis in vivo. Reduction of USP16 led to decreased expression of senescence gene Cdkn2a and mitigated aberrant regulation of the BMP pathway, a previously unknown function of USP16. Thus, we reveal USP16 as a novel target in an AD model that can both ameliorate the NPC defect and rescue memory and learning through its regulation of both Cdkn2a and BMP signaling.'

  View details for DOI 10.7554/eLife.66037

  View details for PubMedID 35311644

• Targeted chromatin ligation, a robust epigenetic profiling technique for small cell numbers. Nucleic acids research Zarnegar, M. A., Reinitz, F. n., Newman, A. M., Clarke, M. F. 2017; 45 (17): e153

  Abstract

  The complexity and inefficiency of chromatin immunoprecipitation strategies restrict their sensitivity and application when examining rare cell populations. We developed a new technique that replaces immunoprecipitation with a simplified chromatin fragmentation and proximity ligation step that eliminates bead purification and washing steps. We present a simple single tube proximity ligation technique, targeted chromatin ligation, that captures histone modification patterns with only 200 cells. Our technique eliminates loss of material and sensitivity due to multiple inefficient steps, while simplifying the workflow to enhance sensitivity and create the potential for novel applications.

  View details for PubMedID 28973448

• An LXR agonist promotes glioblastoma cell death through inhibition of an EGFR/AKT/SREBP-1/LDLR-dependent pathway. Cancer discovery Guo, D., Reinitz, F., Youssef, M., Hong, C., Nathanson, D., Akhavan, D., Kuga, D., Amzajerdi, A. N., Soto, H., Zhu, S., Babic, I., Tanaka, K., Dang, J., Iwanami, A., Gini, B., Dejesus, J., Lisiero, D. D., Huang, T. T., Prins, R. M., Wen, P. Y., Robins, H. I., Prados, M. D., Deangelis, L. M., Mellinghoff, I. K., Mehta, M. P., James, C. D., Chakravarti, A., Cloughesy, T. F., Tontonoz, P., Mischel, P. S. 2011; 1 (5): 442-56

  Abstract

  Glioblastoma (GBM) is the most common malignant primary brain tumor of adults and one of the most lethal of all cancers. Epidermal growth factor receptor (EGFR) mutations (EGFRvIII) and phosphoinositide 3-kinase (PI3K) hyperactivation are common in GBM, promoting tumor growth and survival, including through sterol regulatory element-binding protein 1 (SREBP-1)-dependent lipogenesis. The role of cholesterol metabolism in GBM pathogenesis, its association with EGFR/PI3K signaling, and its potential therapeutic targetability are unknown. In our investigation, studies of GBM cell lines, xenograft models, and GBM clinical samples, including those from patients treated with the EGFR tyrosine kinase inhibitor lapatinib, uncovered an EGFRvIII-activated, PI3K/SREBP-1-dependent tumor survival pathway through the low-density lipoprotein receptor (LDLR). Targeting LDLR with the liver X receptor (LXR) agonist GW3965 caused inducible degrader of LDLR (IDOL)-mediated LDLR degradation and increased expression of the ABCA1 cholesterol efflux transporter, potently promoting tumor cell death in an in vivo GBM model. These results show that EGFRvIII can promote tumor survival through PI3K/SREBP-1-dependent upregulation of LDLR and suggest a role for LXR agonists in the treatment of GBM patients.

  View details for DOI 10.1158/2159-8290.CD-11-0102

  View details for PubMedID 22059152

  View details for PubMedCentralID PMC3207317

• An unbiased screen identifies DEP-1 tumor suppressor as a phosphatase controlling EGFR endocytosis. Current biology : CB Tarcic, G., Boguslavsky, S. K., Wakim, J., Kiuchi, T., Liu, A., Reinitz, F., Nathanson, D., Takahashi, T., Mischel, P. S., Ng, T., Yarden, Y. 2009; 19 (21): 1788-98

  Abstract

  The epidermal growth factor (EGF) stimulates rapid tyrosine phosphorylation of the EGF receptor (EGFR). This event precedes signaling from both the plasma membrane and from endosomes, and it is essential for recruitment of a ubiquitin ligase, CBL, that sorts activated receptors to endosomes and degradation. Because hyperphosphorylation of EGFR is involved in oncogenic pathways, we performed an unbiased screen of small interfering RNA (siRNA) oligonucleotides targeting all human tyrosine phosphatases.We report the identification of PTPRK and PTPRJ (density-enhanced phosphatase-1 [DEP-1]) as EGFR-targeting phosphatases. DEP-1 is a tumor suppressor that dephosphorylates and thereby stabilizes EGFR by hampering its ability to associate with the CBL-GRB2 ubiquitin ligase complex. DEP-1 silencing enhanced tyrosine phosphorylation of endosomal EGFRs and, accordingly, increased cell proliferation. In line with functional interactions, EGFR and DEP-1 form physical associations, and EGFR phosphorylates a substrate-trapping mutant of DEP-1. Interestingly, the interactions of DEP-1 and EGFR are followed by physical segregation: whereas EGFR undergoes endocytosis, DEP-1 remains confined to the cell surface.EGFR and DEP-1 physically interact at the cell surface and maintain bidirectional enzyme-substrate interactions, which are relevant to their respective oncogenic and tumor-suppressive functions. These observations highlight the emerging roles of vesicular trafficking in malignant processes.

  View details for DOI 10.1016/j.cub.2009.09.048

  View details for PubMedID 19836242

  View details for PubMedCentralID PMC2783429