Physical and functional interaction of the p14ARF tumor suppressor with ribosomes

Alterations in the p14(ARF) tumor suppressor are frequent in many human cancers and are associated with susceptibility to melanoma, pancreatic cancer, and nervous system tumors. In addition to its p53-regulatory functions, p14(ARF) has been shown to influence ribosome biogenesis and to regulate the endoribonuclease B23, but there remains considerable controversy about its nucleolar role. We sought to clarify the activities of p14(ARF) by studying its interaction with ribosomes. We show that p14(ARF) and B23 interact within the nucleolar 60 S preribosomal particle and that this interaction does not require rRNA. In contrast to previous reports, we found that expression of p14(ARF) does not significantly alter ribosome biogenesis but inhibits polysome formation and protein translation in vivo. These results suggest a ribosome-dependent p14(ARF) pathway that regulates cell growth and thus complements p53-dependent p14(ARF) functions.     

Rapid micropatterning of cell lines and human pluripotent stem cells on elastomeric membranes

Tissue function during development and in regenerative medicine completely relies on correct cell organization and patterning at micro and macro scales. We describe a rapid method for patterning mammalian cells including human embryonic stem cells (HESCs) and induced pluripotent stem cells (iPSCs) on elastomeric membranes such that micron‐scale control of cell position can be achieved over centimeter‐length scales. Our method employs surface engineering of hydrophobic polydimethylsiloxane (PDMS) membranes by plasma polymerization of allylamine. Deposition of plasma polymerized allylamine (ppAAm) using our methods may be spatially restricted using a micro‐stencil leaving faithful hydrophilic ppAAm patterns. We employed airbrushing to create aerosols which deposit extracellular matrix (ECM) proteins (such as fibronectin and Matrigel?) onto the same patterned ppAAm rich regions. Cell patterns were created with a variety of well characterized cell lines (e.g., NIH‐3T3, C2C12, HL1, BJ6, HESC line HUES7, and HiPSC line IPS2). Individual and multiple cell line patterning were also achieved. Patterning remains faithful for several days and cells are viable and proliferate. To demonstrate the utility of our technique we have patterned cells in a variety of configurations. The ability to rapidly pattern cells at high resolution over macro scales should aid future tissue engineering efforts for regenerative medicine applications and in creating in vitro stem cell niches. Biotechnol. Bioeng. 2012; 109: 2630–2641. © 2012 Wiley Periodicals, Inc.     

The synthesis of E-17 alpha-[131I]iodo-vinyl oestradiol and evaluation of its use as a radiotracer for oestrogen receptor positive breast tumours

17 alpha-Iodo-vinyl oestradiol binds with high affinity to the oestrogen receptor, is metabolically stable and has been shown to accumulate in oestrogen sensitive tissues of rodents. We have synthesised this compound and its 3-acetate, labelled with carrier free 125I and shown the accumulation of both compounds in rat uterus. 17 alpha-Iodo-vinyl oestradiol-3-acetate was prepared labelled with carrier free 131I and administered to twelve patients with suspected breast cancer, approximately 1 h before surgical removal of the primary tumour mass. At the time of surgery a sample of the tumour and a peripheral blood sample were taken. The ratio of radioactivity in the tumour to blood was determined. All tumours accumulated radioactivity and ratios ranged from 1.5:1 to 3.7:1. There was no correlation between the degree of accumulation and either cytosolic oestrogen or progesterone receptor concentration in the tumour. Analysis of blood revealed a single circulating species, 17 alpha-iodo-vinyl oestradiol. The results show that 17 alpha-iodo-vinyl oestradiol is metabolically stable and accumulates in breast tumours though this accumulation is not sufficient to permit imaging.