The dark side of fluorescent protein tagging – the impact of protein tags on biomolecular condensation
Listed in
This article is not in any list yet, why not save it to one of your lists.Abstract
Biomolecular condensation has emerged as an important mechanism to control various cellular processes through the formation of membraneless organelles. Fluorescent protein tags have been extensively used to study the formation and the properties of condensates in vitro and in vivo , but there is evidence that tags may perturb the condensation properties of proteins. In this study, we carefully assess the effects of protein tags on the yeast DEAD-box ATPase Dhh1, a central regulator of processing bodies (P-bodies), which are biomolecular condensates involved in mRNA metabolism. We show that fluorescent tags as well as a poly-histidine tag greatly affect Dhh1 condensation in vitro and lead to condensates with different dynamic properties. Tagging of Dhh1 with various fluorescent proteins in vivo alters the number of P-bodies upon glucose starvation and some tags even show constitutive P-bodies in non-stressed cells. These data raise concerns about the accuracy of tagged protein condensation experiments, highlighting the need for caution when interpreting the results.
Significance Statement
-
Fluorescent tags are extensively used in protein condensation studies although their effect in condensate dynamics has not been carefully investigated.
-
Tags affect the condensation propensity and dynamics of Dhh1 in vitro and P-body numbers in vivo .
-
Tags may generally alter the behavior of proteins in biomolecular condensates and their use needs to be carefully evaluated and controlled.
