How membraneless organelles form
Fluorescent confocal microscopy is revealing why liquid-like droplets made of proteins and DNA form in vitro.
Researchers at the Center for Soft and Living Matter, Institute for Basic Science, South Korea have discovered how membraneless organelles form, in vitro.
The results reveal how DNA sequence is crucial to the formation of these liquid-like droplets of proteins and DNA, shedding light on molecular mechanisms that are closely linked to age-related disease.
To better understand these molecular mechanisms, IBS researchers tested how different sequences of DNA form droplets with a simple protein made from a single repeating amino-acid; lysine (poly-L-lysine).
Analyses revealed that liquid-like droplet formation is related to DNA flexibility and sequence; the more flexible the DNA, the more easily the droplets form.
The researchers discovered that even when the overall electric charge between two DNA molecules is the same, the DNA sequence ultimately determines the stability and appearance of the liquid-like droplets.
"As the rigidity of DNA molecules can be slightly tuned depending on its nucleotide sequence, we could compare DNA molecules with the same change density, but different sequence," explains John King from Microscopy and Spectroscopy at IBS.
Researchers also demonstrated that adenosine triphosphate (ATP), which typically acts as a fuel source in cells, facilitates the formation of liquid-like droplets.
Mixtures of poly-L-lysine and double-stranded DNA, which would typically precipitate at low salt concentrations, readily formed stable liquid-like droplets in presence of ATP.
Fluorescent confocal microscopy image of liquid-like droplets (yellow) formed of poly-L-lysine, DNA (dark spots), and adenosine triphosphate. [IBS]
According to the researchers, this set-up is the perfect platform to examine how the flexibility of nucleic acids affects liquid-liquid phase separation.
"The most fascinating part is to imagine how cells may take advantage of this sequence-dependent information to guide and regulate liquid-liquid phase separation in vivo," concludes King's colleague, Dr Anisha Shakya from IBS.
Research is published in Biophysical Journal.