Significant efforts have been directed to understanding the factors that influence the lineage commitment of stem cells. This paper demonstrates that cell shape, independent of soluble factors, has a strong influence on the differentiation of human mesenchymal stem cells (MSCs) from bone marrow. When exposed to competing soluble differentiation signals, cells cultured in rectangles with increasing aspect ratio and in shapes with pentagonal symmetry but with different subcellular curvature-and with each occupying the same area-display different adipogenesis and osteogenesis profiles. The results reveal that geometric features that increase actomyosin contractility promote osteogenesis and are consistent with in vivo characteristics of the microenvironment of the differentiated cells. Cytoskeletal-disrupting pharmacological agents modulate shape-based trends in lineage commitment verifying the critical role of focal adhesion and myosin-generated contractility during differentiation. Microarray analysis and pathway inhibition studies suggest that contractile cells promote osteogenesis by enhancing c-Jun N-terminal kinase (JNK) and extracellular related kinase (ERK1/2) activation in conjunction with elevated winglesstype (Wnt) signaling. Taken together, this work points to the role that geometric shape cues can play in orchestrating the mechanochemical signals and paracrine/autocrine factors that can direct MSCs to appropriate fates. were initially isolated from bone marrow and noted for their ability to differentiate into bone cells (osteoblasts), cartilage cells (chondrocytes), and fat cells (adipocytes) (1, 2). As an autologous source of stem cells, MSCs are under considerable scrutiny for regenerative therapies (3). A combination of physical, chemical, and biological cues present in the stem cell microenvironment have been implicated as directors of stem cell fate in vivo (4). This concept of a stem cell "niche" has motivated empirical studies to identify optimal combinations of extracellular matrix, culture conditions, and temporally administered growth factors to direct stem cell fate in the laboratory (5-7). However, the physical forces and geometry of the MSC microenvironment remain poorly defined and have begun to emerge as critical parameters for regulating cell fate (8, 9).Physical cues, which were postulated as important factors in tissue development over a century ago (10), have also been recognized as important factors in controlling cell function. Research along these lines has been driven in the past decades with the maturation of microengineering techniques (11-13). Ingber, Whitesides, and colleagues demonstrated the use of substrates that were geometrically patterned to control the microenvironment of individual cells and in turn the decision of cells to initiate apoptosis (14). Patterned substrates have also been used to study cytoskeletal dynamics (15-18) and motility (19-22) with singlecell resolution. Chen and coworkers recently demonstrated the important role that cell shape and size can play ...

In contrast to 5-methylcytosine (5-mC), which has been studied extensively1–3, little is known about 5-hydroxymethylcytosine (5-hmC), a recently identified epigenetic modification present in substantial amounts in certain mammalian cell types4,5. Here we present a method for determining the genome-wide distribution of 5-hmC. We use the T4 bacteriophage β-glucosyltransferase to transfer an engineered glucose moiety containing an azide group onto the hydroxyl group of 5-hmC. The azide group can be chemically modified with biotin for detection, affinity enrichment and sequencing of 5-hmC–containing DNA fragments in mammalian genomes. Using this method, we demonstrate that 5-hmC is present in human cell lines beyond those previously recognized4. We also find a gene expression level–dependent enrichment of intragenic 5-hmC in mouse cerebellum and an age-dependent acquisition of this modification in specific gene bodies linked to neurodegenerative disorders.

Human sex chromosomes evolved from autosomes. Nineteen ancestral autosomal genes persist as differentiated homologs on the X and Y chromosomes. The ages of individual X-Y gene pairs (measured by nucleotide divergence) and the locations of their X members on the X chromosome were found to be highly correlated. Age decreased in stepwise fashion from the distal long arm to the distal short arm in at least four "evolutionary strata." Human sex chromosome evolution was probably punctuated by at least four events, each suppressing X-Y recombination in one stratum, without disturbing gene order on the X chromosome. The first event, which marked the beginnings of X-Y differentiation, occurred about 240 to 320 million years ago, shortly after divergence of the mammalian and avian lineages.