Biology subjects

Unruh, J. R.

Publications and source records attributed to Unruh, J. R..

4 recordsLinked to original sources

bioRxiv2018

Membrane insertion function for SUN-KASH complex revealed by high resolution analysis of yeast centrosomes

Bipolar spindle formation in yeast requires insertion of centrosomes (known as spindle pole bodies (SPBs)) into fenestrated regions of the nuclear envelope (NE). Using structured-illumination microscopy and bimolecular fluorescence complementation, we map protein distribution at SPB fenestra and interrogate protein-protein interactions with high spatial resolution. We find that the Sad1-UNC-84 (SUN) protein Mps3 forms a ring-like structure around the SPB, similar to toroids seen for components of the SPB insertion network (SPIN). Mps3 and the SPIN component Mps2 (a Klarsicht-ANC-1-Syne-1 domain (KASH)-like protein) form a novel non-canonical linker of nucleoskeleton and cytoskeleton (LINC) complex that is connected in both luminal and extraluminal domains. This hairpin-like LINC complex forms during SPB insertion, suggesting it functions in NE reorganization at the pore membrane. The LINC complex also controls the distribution of a soluble SPIN component Bbp1. Taken together our work shows that Mps3 is a fifth SPIN component and suggests both direct and indirect roles for the LINC complex in NE remodeling.

cell biology↗

bioRxiv2018

Synaptonemal complex architecture facilitates the chromosome-specific regulation of recombination in Drosophila

The synaptonemal complex (SC) is a conserved meiotic structure that regulates the repair of double strand breaks (DSBs) into crossovers or gene conversions. The removal of any central region SC component, such as the Drosophila melanogaster transverse filament protein C(3)G, causes a complete loss of SC structure and crossovers. To better understand the role of the SC in meiosis, we used CRISPR/Cas9 to construct three in-frame deletions within the predicted coiled-coil region of the C(3)G protein. These three deletion mutants disrupt SC maintenance at different times during pachytene and exhibit distinct defects in key meiotic processes, allowing us to define the stages of pachytene when the SC is necessary for homolog pairing and recombination. Our studies demonstrate that the X chromosome and the autosomes display substantially different defects in pairing and recombination when SC structure is disrupted, suggesting that the X chromosome is potentially regulated differently than the autosomes.

genetics↗

bioRxiv2017

Quinary structure kinetically controls protein function and dysfunction

Protein self-assemblies modulate protein activities over biological time scales that can exceed the lifetimes of the proteins or even the cells that harbor them. We hypothesized that these time scales relate to kinetic barriers inherent to the nucleation of ordered phases. To investigate nucleation barriers in living cells, we developed Distributed Amphifluoric FRET (DAmFRET). DAmFRET exploits a photoconvertible fluorophore, heterogeneous expression, and large cell numbers to quantify via flow cytometry the extent of a proteins self-assembly as a function of cellular concentration. We show that kinetic barriers limit the nucleation of ordered self-assemblies, and that the persistence of the barriers with respect to concentration relates to structure. Supersaturation resulting from sequence-encoded nucleation barriers gave rise to prion behavior, and enabled a prion-forming protein, Sup35 PrD, to partition into dynamic intracellular condensates or to form toxic aggregates. Our results suggest that nucleation barriers govern cytoplasmic inheritance, subcellular organization, and proteotoxicity.\n\nHighlightsO_LIDistributed Amphifluoric FRET (DAmFRET) quantifies nucleation in living cells\nC_LIO_LIDAmFRET rapidly distinguishes prion-like from non-prion phase transitions\nC_LIO_LINucleation barriers allow switch-like temporal control of protein activity\nC_LIO_LISequence-intrinsic features determine the concentration-dependence of nucleation barriers\nC_LI

biophysics↗

bioRxiv2016

Hoxa1 and TALE proteins display cross-regulatory interactions and form a combinatorial binding code on Hoxa1 targets

Hoxa1 has diverse functional roles in differentiation and development. We have identified and characterized properties of regions bound by Hoxa1 on a genome-wide basis in differentiating mouse ES cells. Hoxa1 bound regions are enriched for clusters of consensus binding motifs for Hox, Pbx and Meis and many display co-occupancy of Pbx and Meis. Pbx and Meis are members of the TALE family and genome-wide analysis of multiple TALE members (Pbx, Meis, TGIF, Prep1 and Prep2) show that nearly all Hoxa1 targets display occupancy of one or more TALE members. The combinatorial binding patterns of TALE proteins defines distinct classes of Hoxa1 targets and indicates a role as cofactors in modulating the specificity of Hox proteins. We also discovered extensive auto- and cross-regulatory interactions among the Hoxa1 and TALE genes. This study provides new insight into a regulatory network involving combinatorial interactions between Hoxa1 and TALE proteins.

developmental biology↗