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The MYC family of oncogenes encodes a set of three related transcription factors that are overexpressed in many human tumors and contribute to the cancer-related deaths of more than 70,000 Americans every year. MYC proteins drive tumorigenesis by interacting with co-factors that enable them to regulate the expression of thousands of genes linked to cell growth, proliferation, metabolism and genome stability. One effective way to identify critical co-factors required for MYC function has been to focus on sequence motifs within MYC that are conserved throughout evolution, on the assumption that their conservation is driven by protein-protein interactions that are vital for MYC activity. In addition to their DNA-binding domains, MYC proteins carry five regions of high sequence conservation known as Myc boxes (Mb). To date, four of the Mb motifs (MbI, MbII, MbIIIa and MbIIIb) have had a molecular function assigned to them, but the precise role of the remaining Mb, MbIV, and the reason for its preservation in vertebrate Myc proteins, is unknown. Here, we show that MbIV is required for the association of MYC with the abundant transcriptional coregulator host cell factor-1 (HCF-1). We show that the invariant core of MbIV resembles the tetrapeptide HCF-binding motif (HBM) found in many HCF-interaction partners, and demonstrate that MYC interacts with HCF-1 in a manner indistinguishable from the prototypical HBM-containing protein VP16. Finally, we show that rationalized point mutations in MYC that disrupt interaction with HCF-1 attenuate the ability of MYC to drive tumorigenesis in mice. Together, these data expose a molecular function for MbIV and indicate that HCF-1 is an important co-factor for MYC.

Lung disease is a major cause of death in the United States, with current therapeutic approaches serving only to manage symptoms. The most common chronic and life-threatening genetic disease of the lung is cystic fibrosis (CF) caused by mutations in the cystic fibrosis transmembrane regulator (CFTR). We have generated induced pluripotent stem cells (iPSCs) from CF patients carrying a homozygous deletion of F508 in the CFTR gene, which results in defective processing of CFTR to the cell membrane. This mutation was precisely corrected using CRISPR to target corrective sequences to the endogenous CFTR genomic locus, in combination with a completely excisable selection system, which significantly improved the efficiency of this correction. The corrected iPSCs were subsequently differentiated to mature airway epithelial cells where recovery of normal CFTR expression and function was demonstrated. This isogenic iPSC-based model system for CF could be adapted for the development of new therapeutic approaches., (Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.)

X-linked Severe Combined Immunodeficiency (SCID-X1) is a genetic disease that leaves newborns at high risk of serious infection and a predicted life span of less than 1 year in the absence of a matched bone marrow donor. The disease pathogenesis is due to mutations in the gene encoding the Interleukin-2 receptor gamma chain (IL-2Rγ), leading to a lack of functional lymphocytes. With the leukemogenic concerns of viral gene therapy there is a need to explore alternative therapeutic options. We have utilized induced pluripotent stem cell (iPSC) technology and genome editing mediated by TALENs to generate isogenic subject-specific mutant and gene-corrected iPSC lines. While the subject-derived mutant iPSCs have the capacity to generate hematopoietic precursors and myeloid cells, only wild-type and gene-corrected iPSCs can additionally generate mature NK cells and T cell precursors expressing the correctly spliced IL-2Rγ. This study highlights the potential for the development of autologous cell therapy for SCID-X1 subjects., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Despite therapeutic advancement, pulmonary disease still remains a major cause of morbidity and mortality around the world. Opportunities to study human lung disease either in vivo or in vitro are currently limited. Using induced pluripotent stem cells (iPSCs), we generated mature multiciliated cells in a functional airway epithelium. Robust multiciliogenesis occurred when notch signaling was inhibited and was confirmed by (i) the assembly of multiple pericentrin-stained centrioles at the apical surface, (ii) expression of transcription factor forkhead box protein J1, and (iii) presence of multiple acetylated tubulin-labeled cilia projections in individual cells. Clara, goblet, and basal cells were all present, confirming the generation of a complete polarized epithelial-cell layer. Additionally, cAMP-activated and cystic fibrosis transmembrane regulator inhibitor 172-sensitive cystic fibrosis transmembrane regulator currents were recorded in isolated epithelial cells. Our report demonstrating the generation of mature multiciliated cells in respiratory epithelium from iPSCs is a significant advance toward modeling a number of human respiratory diseases in vitro.

A tandem mass spectrometric (ms/ms) method using desorption chemical ionization is described for the quantitation of taxol [1], cephalomannine [2], and baccatin III [3] found in Taxus brevifolia bark and needle extracts. A parent ion scan was used to simultaneously determine the weight percentages of 1-3 in bark and needle samples by the method of standard addition. In an alternative experiment, the concentration of 1 in the same samples was determined by ms/ms using trideuterated 10-acetyltaxol [7a] as an internal standard. High-performance liquid chromatography (hplc) was also used to determine the weight percentages of 1-3 in the same T. brevifolia bark and needle extracts with an external standard. The ms/ms method of quantitation by internal standard is the best overall method of analysis examined. With this method, 1 was quantitated in the T. brevifolia extracts at the low picomole level with a relative standard deviation of 17% or better for all samples analyzed with an analysis time of less than five min per sample. The precision, level of quantitation, and speed of analysis of the three methods of taxane quantitation are compared.