Primary Appointment

  Research Interests

• X-ray crystallography
• Transcription initiation
• Transcription regulation
• Partition
• DNA segregation
• Cell division

Research Synopsis
The Schumacher laboratory focuses on understanding, at a detailed atomic level, several key biological processes involving protein-nucleic acid interactions. Our major area of interest is DNA segregation. Ultimately, we wish to obtain structural snapshots of every step of DNA segregation using plasmid partition loci as model systems. In addition, we study several proteins that regulate cohesin removal during anaphase in higher organisms. Finally, we examine several global effectors of transcription in both eukaryotes and prokaryotes to understand their mechanism of transcription activation or repression.

The faithful inheritance of genetic information from parent to offspring is essential for the survival of all cells. Deregulation of this process can have profound effects including the development of aneuploidy, which can lead to cancer. Our goal is to understand the basic molecular principles behind this process. DNA segregation involves the directed movement and positioning of chromosomes, which accurately distributes them to their daughter cells at cell division. This process is mediated by functionally homologous par systems in prokaryotic plasmid systems. The simplicity of plasmid partition systems makes them excellent model systems to address the molecular mechanisms of DNA segregation at a detailed atomic level. Indeed, these systems require only three components: a cis-acting centromere DNA site(s) and two trans-acting proteins, a motor protein and a centromere-binding protein. In the first step of partition, multiple centomere-binding proteins bind cooperatively to the centromere-site, which generally consists of several tandem repeats, to form a higher order protein-DNA structure called a segrosome. This structure serves as the assembly site for the motor protein, which then mediates DNA separation of paired, replicated plasmids. There are two major par systems, those that use actin like motor proteins and those that employ Walker type ATPase proteins. We are examining partition by both systems, including the P1 plasmid par system and the multidrug resistant pSK41 par system. Our studies on the P1 system have provided structures of the centomere-binding protein, ParB, bound to minimal centromere elements. These structures have revealed that ParB is a novel DNA-binding protein. Its bridging and pairing capabilities explain how it binds its complex centromere and mediate plasmid pairing. We also recently obtained the first structure of a segrosome, that of the pSK41 ParR-centomere complex. This structure showed that the segrosome is a very large superhelical protein-nucleic acid structure with dimensions ideal for capturing the actin-like filaments recently shown to be formed by the motor ATPase protein. Our structures and biochemical analyses on both centromere-binding and motor proteins and their complexes will provide the foundation for understanding basic molecular mechanisms of how proteins and multiprotein-DNA complexes such as the segrosome, function to mediate and drive plasmid/chromosome segregation.

In our studies of transcription one area of interest is the mechanism of transcription initiation by the primitive eukaryote, T. vaginalis. The transcription start sites of nearly all T. vaginalis genes contain an Inr element that appears to be primarily responsible for start site selection and is recognized by one protein, the Initiator Binding Protein, 39 kDa, IBP39. Thus, compared to higher eukaryotes, T. vaginalis, which is considered the earliest extant eukaryote, provides a simplified model system to study transcription start site selection. IBP39 shows no sequence similarity to any protein and consists of a N-terminal, 14.5 kDa, Inr binding domain (IBD) connected by a long, flexible linker to a C-domain of previously unknown function. Our structural and biochemical studies revealed the basis for recognition of the short and loose consensus Inr sequence by IBP39 and also demonstrated that the C-domain is involved in recruitment of the RNA polymerase II C-terminal domain (CTD). This recruitment likely explains its role in mediating transcription initiation. Subsequent structural studies aimed at obtaining multi-protein core promoter complexes that possibly include the T. vaginalis RNA polymerase II are envisioned.

Last updated: 5/29/2008