| RESEARCH HIGHLIGHTS |


two novel regulators of cell fate — all of which was validated with wet lab experiments. The researchers also considered the process of


human muscle cell development, demonstrating the intricate crosstalk between muscle and non-muscle cell lineages and revealing seven waves of gene expression changes during differentiation. According to Chen, “Mpath can


be used to trace lineages of any cell type.” For example, one could extend the method to study “how cancer cells have progressed from benign to malignant, as well as which genes are driving the progression,” she says. Chen and her colleagues are now working


on an enhanced version of the algorithm called Mtree, which would allow the simultaneous


analysis of two or more independent development pathways, such as in the brain where different immune lineages are distinct from other tissues.


1. Chen, J., Schlitzer, A., Chakarov, S., Ginhoux, F. & Poidinger, M. Mpath maps multi-branching single-cell trajectories revealing progenitor cell progression during development. Nature Communications 7, 11988 (2016).


Fluorescence imaging:


CELLULAR THERMOMETER


LIGHT-EMITTING DYES CAN MEASURE HEAT GENERATION WITHIN LIVING CELLS WITH SUBCELLULAR ACCURACY


A technique that uses fluorescent dyes to measure the temperature inside living cells is helping to reveal the mechanism by which living organisms generate heat. An international team co-led by A*STAR


researchers have shown that the dyes, which adjust their light emission in response to tem- perature, can be used to measure heat generation within muscle cells. The technique has already resolved one controversy over the amount of heat such cells can generate. Warm-blooded animals must produce a


significant amount of heat to maintain their temperature. Muscle tissue is one source, generating heat through shivering. But muscle cells can also heat up via a chemical process called non-shivering thermogenesis (NST). This process is less well understood, and even the esti- mates of the amount of heat generated this way by the body can differ wildly. Some tests have suggested they could raise their temperature by


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Fluorescence lifetime imaged on a color scale — at higher tempera- tures (as above), the fluorescent dye loses its fluorescence more rapidly, which can be detected to measure temperature change.


one degree Celsius via NST, whereas an estimate calculated from a mass of cells suggested a mere 10—5


degree Celsius per cell. Birgitte Lane and Hideki Itoh at the A*STAR


Institute of Medical Biology, along with collab- orators in Japan, have unraveled these results using temperature-responsive dyes developed at the Singapore Bioimaging Consortium. The team used ER thermo yellow, a dye that


sticks to the structure in muscle cells thought to be the site of NST, the sarcoplasmic reticulum. The dye fluoresces after being exposed to light, and the length of time it remains illuminated varies with temperature. Using A*STAR’s analytical fluorescence


microscopy facility, the researchers could measured the fluorescent lifetime of the dye with sub-nanosecond accuracy. When they dosed muscle cells with caffeine, they detected a temperature increase of around 1.6 degrees Celsius using ER thermo yellow. When they


repeated the experiment using another dye that diffuses throughout the cell, no such tempera- ture increase was observed, confirming the heat was generated at the sarcoplasmic reticulum. “Using this approach allows us to look at heat


generation much more accurately within cells, so we can see which cell types have this capability and where in the cell the heat generation is taking place. We can then start to dissect the mechanism,” says Lane. The result confirms that specialized cells


within muscle tissue are able to raise their temperature by around 1 degree Celsius via NST. This new methodology could now be used for medical applications such as screening drug for conditions like obesity or heat regulation disorders like malignant hypothermia.


1. Itoh, H., Arai, S., Sudhaharan, T., Lee, S.-C., Chang, Y.-T., et al. Direct organelle thermometry with fluorescence lifetime imaging microscopy in single myotubes. Chemical Communications 52, 4458–4461 (2016).


A*STAR RESEARCH 29


Reproduced from Ref. 1 by permission of the Royal Society of Chemistryw


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