Research in the Wuttke lab spans two main areas, telomere biology and plasticity in binding. Details on projects in these areas can be found at our lab web page.
Telomere Biology
Telomeres are specialized nucleoprotein structures at the ends of eukaryotic chromosomes that are essential for chromosome stability and cellular proliferation. Telomeric DNA does not encode for proteins, instead it consists of tandem repeats of TG-rich sequences of double-stranded DNA that terminate in a 3¢ single-stranded DNA overhang. Protection of this overhang is essential. When left unprotected, this overhang initiates DNA damage responses that lead to catastrophic events permanently damaging the genome and resulting in apoptosis or senescence. Furthermore, telomere shortening due to the inability of the DNA-replication machinery to fully replicate the ends is a critical mechanism of tumor suppression as well as a hallmark of aging. Continually proliferating cells maintain adequate telomeres through the action of the reverse transcriptase telomerase.Telomeres are important to human health because dysregulation of either telomere protection or telomerase activity causes many human diseases. Notably, over 90% of human cancers activate telomerase for continued proliferation.
Our research in this area aims to understand how telomere-associated proteins protect and maintain telomeres. Key questions include how subunits of the telomerase enzyme contribute to activity, how the single-strand DNA overhang is shielded from the DNA-damage machinery, and whether capping activity also regulates telomerase action. We develop this knowledge by first understanding the core activities of key telomere factors, then testing these activities in a reconstituted telomerase assay and validating our knowledge directly in the organism.
Plasticity in Molecular Recognition
Many biologically critical recognition events involve the specific binding of flexible ligands such as single-stranded (ss) DNA, RNA, peptides and carbohydrates. Structural plasticity, defined as the ability of an interface to adopt alternate conformations when bound to different ligands, has been invoked to explain binding specificity and promiscuity in several protein/ligand systems. Furthermore, an understanding of the malleability of a binding interface is increasingly recognized as key to predicting its binding activity and specificity. Discerning the scope and mechanisms of rearrangements at binding interfaces is essential to understanding the biophysics of molecular recognition events. The focus of this proposal is to investigate the extent of structural plasticity in the recognition of these flexible ligands.
We use the recognition of ssDNA by the telomere end-binding proteins as the predominant model to characterize the contribution of structural plasticity to recognition. The telomere-end binding proteins Pot1 and Cdc13 bind the conserved 3’ ssDNA overhang at telomeres. This binding is required for cellular viability. However, the sequence of the overhang is somewhat variable, meaning that these proteins need to bind divergent ligands while maintaining exquisite specificity. Extensive evidence suggests that the protein/nucleic acid interface adopts altered configurations in the presence of different ligands that bind with similar affinities. We are investigating the hypothesis that this structural plasticity is important for specificity. Moreover, the malleability of the interface may further contribute to function by providing a way to physically alter the structure and accessibility of the 3’ end. We us an integrated set of strategies to address this question, ranging from determination of high-resolution structures to in vivo assessment of activities.
