Integrated Calendar

Our organs and tissues are made of different cell types that communicate with each other in order to achieve joint functions. However, little is known about the universal principles of these interactions. For example, how do cell interactions maintain stable cell composition, spatial organization and collective division of labor in tissues?
And what is the role of these interactions in tissue-level diseases where the healthy balance in the tissue is disrupted such as excess scarring following injury known as fibrosis? In this talk, I will discuss my work in developing new theoretical frameworks that explore the collective behavior of cells that emerges from cell-cell interactions.
I will present work on the cell communication circuit that controls tissue repair following injury and how it may lead to fibrosis. I will discuss a new mathematical approach to explore how cell interactions can be used to provide symmetry breaking and optimal division of labor in tissues, and how this approach can help to interpret complex patterns in real high-dimensional data.
I will introduce a new concept in complex networks – network hyper-motifs, where we explore how small recurring patterns (network motifs) are integrated within large networks, and how these larger patterns (hyper-motifs) can give rise to emergent dynamic properties. Finally, I will conclude with future directions that are aimed at revealing principles that unify our understanding of different tissues.

Many peptides and proteins form amyloid fibrils whose detailed molecular structure is of
considerable functional and pathological importance. For example, amyloid is closely associated
with the neurodegenerative diseases such as Alzheimer’s and Parkinson’s diseases. We review the
macroscopic properties of fibrils and outline approaches to determining their microscopic structure
using magic angle spinning (MAS) NMR with 2D and 3D dipole recoupling experiments involving
spectral assignments and distance measurements. Key to obtaining high resolution is measurement
of a sufficient number of NMR structural restraints (13C-13C and 13C-15N distances per residue). In
addition, we demonstrate the applicability of 1H detection and dynamic nuclear polarization (DNP)
to amyloid structural studies.
We discuss the structures of three different amyloids: (1) fibrils formed by Ab1-42, the toxic
species in Alzheimer’s, using >500 distance constraints; (2) fibrils of Ab1-40, a second form of Ab
with a different structure, and (3) a structure of fibrils forned by b2-microglobulin, the 99 amino
acid protein associated with dialysis related amylosis, using ~1200 constraints. Contrary to
conventional wisdom, the spectral data indicate that the molecules in the fibril are microscopically
well ordered. In addition, the structures provide insight into the mechanism of interaction of the
monoclonal antibody, Aducanumab, directed against Ab amyloid.

Dynamic nuclear polarization (DNP) has become an invaluable tool to enhance sensitivity of
magic angle spinning (MAS) NMR, enabling the study of biomolecules and materials which are
otherwise intractable. In this presentation we explore some new aspects of time domain DNP
experiments and their applications.
One of the main thrusts of DNP was to provide increased sensitivity for MAS spectroscopy of
membrane and amyloid protein experiments. A problem frequently encountered in these
experiments is the broadened resonances that occur at low temperatures when motion is quenched.
In some cases it is clear that the proteins are homogeneously broadened, and therefore that higher
Zeeman fields and faster spinning is required to recall the resolution. We show this is the case for
MAS DNP spectra of Ab1-42 amyloid fibrils where the resolution at 100 K is identical to that at room
temperature. Furthermore, we compare the sensitivity of DNP and 1H detected experiments and find
that DNP, even with a modest ℇ=22, is ~x6.5 times more sensitive.
We have also investigated the frequency swept-integrated solid effect (FS-ISE) and two recently
discovered variants – the stretched solid effect (SSE) and the adiabatic solid effect (ASE). We find
that the latter two experiments can give up to a factor of ~2 larger enhancement than the FS-ISE.
The SSE and ASE experiments should function well at high fields.
Finally, we discuss two new instrumental advances. First, a frequency swept microwave source
that permits facile investigation of field profiles. It circumvents the need for a B0 sweep coil and the
compromise of field homogeneity and loss of helium associated with such studies. This
instrumentation has permitted us to elucidate the polarization transfer mechanism of the Overhauser
effect, and also revealed interesting additional couplings (ripples) in field profiles of cross effect
polarizing agents. Second, to improve the spinning frequency in DNP experiments, we have
developed MAS rotors laser machined from single crystal diamonds. Diamond rotors should permit
higher spinning frequencies, improved microwave penetration, and sample cooling.

For many proteins, flexibility and motion form the basis of their function. In our lab, we quantify the conformational landscapes of proteins and their changes upon interaction with external effectors. Using Double Electron-Electron Resonance (DEER) spectroscopy, a form of Electron Paramagnetic Resonance (EPR) spectroscopy, we directly measure absolute distances and distance distributions between pairs of spin labels within proteins. From the data, we build quantitative structural and energetic models of the protein's intrinsic flexibility, conformational substates, and the structural changes induced by ligands and binding partners. In this talk, I present some of our recent results on the allosteric regulation of ion channels, the function of de novo designed protein switches, and novel methods for measuring protein conformations directly in their native cellular environment.