Pyrin - A Key Regulator of Inflammation
Sudarshan Seshadri
IL-1β is a
proinflammatory cytokine, which plays an important role in innate host
defense and host homeostasis. It is synthesized as a pro-form
(proIL-1β) which gets processed to mature
IL-1β by an enzyme known as caspase-1. Caspase-1 is also
synthesized as a precursor protein, which requires activation by a
multiprotein complex known as the inflammasome. Apoptotic speck like
protein containing a CARD domain (ASC) is an integral adaptor protein
of the inflammasome and is also an activator of caspase-1. Macrophages
and their precursors monocytes, are the key cells and form the
frontline of defense in the innate immune response. Although both
monocytes and macrophages express caspase-1, macrophages are limited
in their ability to activate the enzyme and release functional
IL-1β. Since, mutations in the pyrin gene (MEFV) cause
the inflammatory disorder familial Mediterranean fever (FMF), we
hypothesized that pyrin regulates caspase-1 activation by interacting
with ASC and caspase-1. In order to determine the whether variations
in pyrin expression explain the difference between monocytes and
macrophages in IL-1β processing and release, pyrin was studied in
human monocytes and monocytes derived macrophages. While monocytes
readily express pyrin mRNA and protein upon endotoxin stimulation,
macrophages express significantly less pyrin. Pyrin levels directly
correlated with IL-1β processing and release, therefore we asked
whether pyrin may augment IL-1β processing and
release. Overexpression of pyrin induced IL-1β processing
and release in a dose dependent fashion in HEK293 cells. Knock down of
pyrin by using small interfering RNA suppressed IL-1β
processing and release in both THP-1 cells and human monocytes. Our
data suggests that pyrin is one of the key positive regulators of
IL-1β processing and release in monocytes and plays an
important role in endotoxin induced inflammation. Understanding the
role of pyrin and pyrin mutants will be helpful in understanding
mechanisms of inflammation caused by FMF and may open new avenues for
therapeutic intervention in FMF.
Solution NMR of Large Molecules and Assemblies
Mark P. Foster, Craig A. McElroy, and Carlos D.
Amero
Solution NMR spectroscopy represents a powerful tool for examining the structure and function of biological macromolecules. The advent of multidimensional (2D-4D) NMR, together with the widespread use of uniform isotopic labeling of proteins and RNA with the NMR-active isotopes, 15N and 13C, opened the door to detailed analyses of macromolecular structure, dynamics, and interactions of smaller macromolecules (<~25 kDa). Over the past 10 years, advances in NMR and isotope labeling methods have expanded the range of NMR-tractable targets by at least an order of magnitude. Here we briefly describe the methodological advances that allow NMR spectroscopy of large macromolecules and their complexes and provide a perspective on the wide range of applications of NMR to biochemical problems.