Unblending of Transcriptional Condensates in Human Repeat Expansion Disease

Genetic disorders occur when a person’s DNA sequence is altered away from the normal sequence. This can result from the mutation of one or multiple genes, environmental factors in conjunction with gene mutations, or chromosomal damage. Many genetic disorders are inherited at birth. Of those, more than 30 are caused by an abnormal expansion of short, repetitive DNA sequence elements. Most of these repeat expansions result in expansions of alanine or glutamine in cellular proteins which are linked to developmental disorders and neurodegenerative diseases, respectively. Previous studies have not focused on the impacts of repeat expansions on protein function but rather on tendencies to form abnormal aggregates, and changes in subcellular localization or proteolytic processing. However, understanding the effects of repeat expansions on the function of impacted proteins is essential for determining how these diseases can potentially be treated.

In this paper from Basu et al. in Cell, the authors investigated repeat expansions occurring in intrinsically disordered regions (IDRs) of transcription factors (TFs) and their effects on the TFs phase separation capacity and ability to form transcriptional condensates. They found that disease-associated repeat expansions in the studied TFs resulted in altered phase separation capacity and ability to co-condense with transcriptional co-activators in comparison to wildtype TFs, resulting in the proposal that changes to TF phase separation and condensate formation may play an important role in human pathologies associated with repeat expansions of TF IDRs. The Vutara 352 imaging platform along with SRX software was utilized for localization, visualization, and statistical analysis of STORM data (stochastic optical reconstruction microscopy). Authors imaged wildtype (HOXD13) and mutated protein (HOXD13 + 7A alleles) and a co-stain of BRD4, a co-activator, to identify that HOXD13-condensates have altered composition in vivo. To observe this, they used co-localization analysis in SRX to determine the Manders overlap coefficients of HOXD13 with BRD4 and HOXD13 +7A with BRD4 and found that wildtype HOXD13 puncta had a higher degree of colocalization with BRD4 than the repeat expansion variant, indicating alteration in condensate composition as a result of the repeat expansions. Super-resolution microscopy was necessary to make this observation as the HOXD13 puncta are less than 100 nm in size, well below the diffraction limit of light.