SummaryHuman pluripotent stem cells (hPSCs) maintain a highly fragmented mitochondrial network, but the mechanisms regulating this phenotype remain unknown. Here, we describe a non-cell death function of the anti-apoptotic protein, MCL-1, in regulating mitochondrial dynamics and promoting pluripotency of stem cells. MCL-1 is induced upon reprogramming, and its inhibition or knockdown induces dramatic changes to the mitochondrial network as well as loss of the key pluripotency transcription factors, NANOG and OCT4. Aside from localizing at the outer mitochondrial membrane like other BCL-2 family members, MCL-1 is unique in that it also resides at the mitochondrial matrix in pluripotent stem cells. Mechanistically, we find MCL-1 to interact with DRP-1 and OPA1, two GTPases responsible for remodeling the mitochondrial network. Depletion of MCL-1 compromised the levels and activity of these key regulators of mitochondrial dynamics. Our findings uncover an unexpected, non-apoptotic function for MCL-1 in the maintenance of mitochondrial structure and stemness.

Advances in single-cell biology have enabled measurements of >40 protein features on millions of immune cells within clinical samples. However, the data analysis steps following cell population identification are susceptible to bias, time-consuming, and challenging to compare across studies. Here, an ensemble of unsupervised tools was developed to evaluate four essential types of immune cell information, incorporate changes over time, and address diverse immune monitoring challenges. The four complementary properties characterized were: 1) systemic plasticity, 2) change in population abundance, 3) change in signature population features, and 4) novelty of cellular phenotype. Three systems immune monitoring studies were selected to challenge this ensemble approach. In serial biopsies of melanoma tumors undergoing targeted therapy, the ensemble approach revealed enrichment of double-negative (DN) T cells. Melanoma tumor resident DN T cells were abnormal and phenotypically distinct from those found in non-malignant lymphoid tissues, but similar to those found in glioblastoma and renal cell carcinoma. Overall, ensemble systems immune monitoring provided a robust, quantitative view of changes in both the system and cell subsets, allowed for transparent review by human experts, and revealed abnormal immune cells present across multiple human tumor types.

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The core transcriptional network regulating stem cell self-renewal and pluripotency remains an intense area of research. Increasing evidence indicates that modified regulation of basic cellular processes such as mitochondrial dynamics, apoptosis, and cell cycle are also essential for pluripotent stem cell identity and fate decisions. Here, we review evidence for Wnt regulation of pluripotency and self-renewal, and its connections to emerging features of pluripotent stem cells, including (1) increased mitochondrial fragmentation, (2) increased sensitivity to cell death, and (3) shortened cell cycle. We provide a general overview of the stem cell–specific mechanisms involved in the maintenance of these uncharacterized hallmarks of pluripotency and highlight potential links to the Wnt signaling pathway. Given the physiological importance of stem cells and their enormous potential for regenerative medicine, understanding fundamental mechanisms mediating the crosstalk between Wnt, organelle-dynamics, apoptosis, and cell cycle will be crucial to gain insight into the regulation of stemness.