RESEARCH

SINGLE MOLECULE PROTEIN-DNA INTERACTIONS

Characterization of DNA binding sites for transcription factors and other proteins is of fundamental importance in molecular biology. It is commonly addressed experimentally by methods like chromatin immunoprecipitation and sequencing (ChIP-seq) of bulk samples. We have developed an alternative method called Chromatin Antibody-mediated Methylating Protein (ChAMP). It is comprised of a GpC methyltransferase fused to protein G. By tethering ChAMP to a primary antibody directed against the DNA-binding protein of interest, and selectively switching on its enzymatic activity in situ, we generated distinct and identifiable methylation patterns next to the protein binding sites. By passing long single molecules through a nanopore, we are able to detect where the protein of interest interacted with the molecule. We aim to increase the capabilities of this technology, multiplexing it to detect the interactions of many distinct proteins on a single DNA molecule. The successful implementation of this technology will enable us to answer fundamental questions about DNA regulation, epigenetics and translation.

Epigenetic information loss is a common feature of aging and multiple diseases

Two seemingly unrelated questions in the field of aging are “Is aging a disease?” and the role of epigenetic information loss during aging. While the first is often viewed as almost a philosophical question, the second concerns the exact roles of a well established driver of aging. We demonstrate that tissue-specific DNA methylation patterns, a form of epigenetic information, are lost during normal aging. Furthermore, we reveal that this information loss is a shared characteristic of various diseases, primarily affecting the diseased organ. This suggests that, from an information-loss standpoint, aging can be considered a disease. Moreover, several diseases can be viewed as instances of organ-specific aging.

SCREENING FOR PATHOGENIC AND AUTO ANTIBODIES THAT MODULATE THE RISK FOR ALZHEIMER’S DISEASE

Alzheimer’s disease (AD) has been associated with infections generated by several pathogens such as Herpes simplex virus 1 (HSV-1) and the periodontopathic bacterium Porphyromonas gingivalis. P. gingivalis have been demonstrated in brain samples of AD patients and in mice and cellular models was linked to the production of a key characteristic of AD- the amyloid β (Aβ) peptide (the major component of the amyloid plaque). On the other hand, blocking P. gingivalis neurotoxicity by small molecules inhibits its virulence factors, reduces neuroinflammation and preserves neural cells in the hippocampus. Interestingly, though HSV-1/P. gingivalis exposure is very prevalent, only a portion of affected individuals will develop AD. We propose that specific immune responses modulate the risk of developing AD following exposure to pathogens. We test this hypothesis by screening saliva and blood samples of nondemented elderly individuals (No-AD) for antibodies against AD-linked pathogens and for autoantibodies for brain (normal and AD) proteins. The immune response of No-AD, AD patients (AD) and younger controls (C) to various pathogens will be compared. A successful identification of a protective immune response will strengthen the pathogenic link to AD, provide an invaluable early diagnostic tool and establish a path toward development of a vaccine.

CHARACTERIZATION OF STREPTOCOCCUS MUTANS ANTIGENS INDUCING SECRETORY IMMUNOGLOBULIN A ANTIBODIES IN HEALTHY CHILDREN

Childhood caries is an ongoing epidemic, resulting in many dental interventions during childhood and adulthood. Dental plaque and more specifically Streptococcus mutans (S. mutans), have a major role in this multifactorial disease. We will characterize the different immunologic response to S.mutans by Secretory Immunoglobulin A (SIgA) and associate it to Decayed, Missing, and Filled Permanent Teeth (DMFT) score. The reportuare of antigen identified by the immune system of children and adolescents with low and high DMFT scores will be characterized and compared. Antibodies from unstimulated saliva will be extracted, exposed to S. mutans protein lysate and analysed by mass-spectrometry. Proteins preferentially bound by the low DMFT group may suggest a caries-protective role for antibodies to that protein enable early identification of children with elevated risk of caries.