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Baryawno, Ninib

The DNA repair enzyme O6-methylguanine-DNA methyltransferase (MGMT) is commonly overexpressed in cancers and is implicated in the development of chemoresistance. The use of drugs inhibiting MGMT has been hindered by their haematologic toxicity and inefficiency. As a different strategy to inhibit MGMT we investigated cellular regulators of MGMT expression in multiple cancers. Here we show a significant correlation between Wnt signalling and MGMT expression in cancers with different origin and confirm the findings by bioinformatic analysis and immunofluorescence. We demonstrate Wnt-dependent MGMT gene expression and cellular co-localization between active β-catenin and MGMT. Pharmacological or genetic inhibition of Wnt activity downregulates MGMT expression and restores chemosensitivity of DNA-alkylating drugs in mouse models. These findings have potential therapeutic implications for chemoresistant cancers, especially of brain tumours where the use of temozolomide is frequently used in treatment.

The mTOR pathway is a critical determinant of cell persistence and growth wherein mTOR complex 1 (mTORC1) mediates a balance between growth factor stimuli and nutrient availability. Amino acids or glucose facilitates mTORC1 activation by inducing RagA GTPase recruitment of mTORC1 to the lysosomal outer surface, enabling activation of mTOR by the Ras homolog Rheb. Thereby, RagA alters mTORC1-driven growth in times of nutrient abundance or scarcity. Here, we have evaluated differential nutrient-sensing dependence through RagA and mTORC1 in hematopoietic progenitors, which dynamically drive mature cell production, and hematopoietic stem cells (HSC), which provide a quiescent cellular reserve.In nutrient-abundant conditions, RagA-deficient HSC were functionally unimpaired and upregulated mTORC1 via nutrientinsensitive mechanisms. RagA was also dispensable for HSC function under nutritional stress conditions. Similarly, hyperactivation of RagA did not affect HSC function. In contrast, RagA deficiency markedly altered progenitor population function and mature cell output. Therefore, RagA is a molecular mechanism that distinguishes the functional attributes of reactive progenitors from a reserve stem cell pool. The indifference of HSC to nutrient sensing through RagA contributes to their molecular resilience to nutritional stress, a characteristic that is relevant to organismal viability in evolution and in modern HSC transplantation approaches.

Recent single-cell RNA-seq protocols based on droplet microfluidics use massively multiplexed barcoding to enable simultaneous measurements of transcriptomes for thousands of individual cells. The increasing complexity of such data creates challenges for subsequent computational processing and troubleshooting of these experiments, with few software options currently available. Here, we describe a flexible pipeline for processing droplet-based transcriptome data that implements barcode corrections, classification of cell quality, and diagnostic information about the droplet libraries. We introduce advanced methods for correcting composition bias and sequencing errors affecting cellular and molecular barcodes to provide more accurate estimates of molecular counts in individual cells. Electronic supplementary material The online version of this article (10.1186/s13059-018-1449-6) contains supplementary material, which is available to authorized users.