5 research outputs found
Click Quantitative Mass Spectrometry Identifies PIWIL3 as a Mechanistic Target of RNA Interference Activator Enoxacin in Cancer Cells
Enoxacin
is a small molecule that stimulates RNA interference (RNAi)
and acts as a growth inhibitor selectively in cancer but not in untransformed
cells. Here, we used alkenox, a clickable enoxacin surrogate, coupled
with quantitative mass spectrometry, to identify PIWIL3 as a mechanistic
target of enoxacin. PIWIL3 is an Argonaute protein of the PIWI subfamily
that is mainly expressed in the germline and that mediates RNAi through
piRNAs. Our results suggest that cancer cells re-express PIWIL3 to
repress RNAi through miRNAs and thus open a new opportunity for cancer-specific
targeting
Alternative Routes to Tricyclic Cyclohexenes with Trinuclear Palladium Complexes
Highly
symmetric all-metal aromatic Pd<sub>3</sub><sup>+</sup> complexes
can catalyze the cycloisomerization of terminal 1,6-enynes and internal
dienynes under mild conditions. Modification of substrates dictates
the mechanism and steers the reaction toward different polycyclic
frameworks, enabling the development of complex cascades. The reactivity
of Pd(<sup>4</sup>/<sub>3</sub>) complexes is complementary to that
of mononuclear Pd(0) and Pd(II) ones
Pd Catalysis in Cyanide-Free Synthesis of Nitriles from Haloarenes via Isoxazolines
A method to obtain
aryl nitriles from the corresponding halides
by Pd catalysis, in the absence of any cyanide source, is reported.
The reaction of an aryl halide, ethyl nitroacetate, and an olefin
readily delivers an aromatic nitrile. A variety of aryl iodides/bromides
have been converted into the corresponding cyanoarenes in fair to
excellent yields. The reaction likely involves the following steps:
(a) Pd-catalyzed α-arylation of ethyl nitroacetate; (b) nitrile
oxide formation; (c) [3 + 2]-cycloaddition with an olefin to provide
an isoxazoline; (d) isoxazoline cleavage to benzonitrile formation
Nonlinear Emission of Quinolizinium-Based Dyes with Application in Fluorescence Lifetime Imaging
Charged
molecules based on the quinolizinum cation have potential
applications as labels in fluorescence imaging in biological media
under nonlinear excitation. A systematic study of the linear and nonlinear
photophysics of derivatives of the quinolizinum cation substituted
by either dimethylaniline or methoxyphenyl electron donors is performed.
The effects of donor strength, conjugation length, and symmetry in
the two-photon emission efficiency are analyzed in detail. The best
performing nonlinear fluorophore, with two-photon absorption cross
sections of 1140 GM and an emission quantum yield of 0.22, is characterized
by a symmetric D-π-A<sup>+</sup>-π-D architecture based
on the methoxyphenyl substituent. Application of this molecule as
a fluorescent marker in optical microscopy of living cells revealed
that, under favorable conditions, the fluorophore can be localized
in the cytoplasmatic compartment of the cell, staining vesicular shape
organelles. At higher dye concentrations and longer staining times,
the fluorophore can also penetrate into the nucleus. The nonlinearly
excited fluorescence lifetime imaging shows that the fluorophore lifetime
is sensitive to its location in the different cell compartments. Using
fluorescence lifetime microscopy, a multicolor map of the cell is
drafted with a single dye
Family-wide Analysis of the Inhibition of Arf Guanine Nucleotide Exchange Factors with Small Molecules: Evidence of Unique Inhibitory Profiles
Arf
GTPases and their guanine nucleotide exchange factors (ArfGEFs)
are major regulators of membrane traffic and organelle structure in
cells. They are associated with a variety of diseases and are thus
attractive therapeutic targets for inhibition by small molecules.
Several inhibitors of unrelated chemical structures have been discovered,
which have shown their potential in dissecting molecular pathways
and blocking disease-related functions. However, their specificity
across the ArfGEF family has remained elusive. Importantly, inhibitory
responses in the context of membranes, which are critical determinants
of Arf and ArfGEF cellular functions, have not been investigated.
Here, we compare the efficiency and specificity of four structurally
distinct ArfGEF inhibitors, Brefeldin A, SecinH3, M-COPA, and NAV-2729,
toward six ArfGEFs (human ARNO, EFA6, BIG1, and BRAG2 and Legionella and Rickettsia RalF). Inhibition was assessed by fluorescence kinetics using pure
proteins, and its modulation by membranes was determined with lipidated
GTPases in the presence of liposomes. Our analysis shows that despite
the intra-ArfGEF family resemblance, each inhibitor has a specific
inhibitory profile. Notably, M-COPA is a potent pan-ArfGEF inhibitor,
and NAV-2729 inhibits all GEFs, the strongest effects being against
BRAG2 and Arf1. Furthermore, the presence of the membrane-binding
domain in Legionella RalF reveals a
strong inhibitory effect of BFA that is not measured on its GEF domain
alone. This study demonstrates the value of family-wide assays with
incorporation of membranes, and it should enable accurate dissection
of Arf pathways by these inhibitors to best guide their use and development
as therapeutic agents