Streamlined histone-based fluorescence lifetime imaging microscopy (FLIM) for studying chromatin organisation.

Changes in chromatin structure are key determinants of genomic responses. Thus, methods that enable such measurements are instrumental for investigating genome regulation and function. Here, we report further developments and validation of a streamlined method of histone-based fluorescence lifetime imaging microscopy (FLIM) that robustly detects chromatin compaction states in fixed and live cells, in 2D and 3D.

Both chondroinduction and proliferation account for growth of cartilage nodules in mouse limb bud cultures.

High density micromass culture of limb bud mesenchymal stem cells isolated from mouse embryos represents a well-established model to study chondro- and osteogenesis. In spite of wide usage of the limb bud model, the mechanisms underlying cartilage nodule growth remain unclear. To determine whether cartilage nodules grow solely by induction of surrounding cells or proliferation of cells within the nodules, we performed BrdU/Collagen II (Col II) double-labelling and 3D reconstruction of growing cartilage nodules.

Calreticulin and focal-contact-dependent adhesion.

Cell adhesion is regulated by a variety of Ca2+-regulated pathways that depend on Ca2+-binding proteins. One such protein is calreticulin, an ER-resident protein. Calreticulin signalling from within the ER can affect processes outside the ER, such as expression of several adhesion-related genes, most notably vinculin and fibronectin.