Neural-specific deletion of Htra2 causes cerebellar neurodegeneration and defective processing of mitochondrial OPA1.

HTRA2, a serine protease in the intermembrane space, has important functions in mitochondrial stress signaling while its abnormal activity may contribute to the development of Parkinson's disease. Mice with a missense or null mutation of Htra2 fail to thrive, suffer striatal neuronal loss, and a parkinsonian phenotype that leads to death at 30-40 days of age. While informative, these mouse models cannot separate neural contributions from systemic effects due to the complex phenotypes of HTRA2 deficiency.

Protoporphyrins enhance oligomerization and enzymatic activity of HtrA1 serine protease.

High temperature requirement protein A1 (HtrA1), a secreted serine protease of the HtrA family, is associated with a multitude of human diseases. However, the exact functions of HtrA1 in these diseases remain poorly understood. We seek to unravel the mechanisms of HtrA1 by elucidating its interactions with chemical or biological modulators.

Microtubule dynamics regulates Akt signaling via dynactin p150.

Following activation at the plasma membrane, Akt is subsequently deactivated in the cytoplasm. Although activation and deactivation of Akt must sometimes be separated in order to elicit and control cellular responses, the exact details of the spatiotemporal organization of Akt signaling are incompletely understood. Here we show that microtubule dynamics specifically modulate the deactivation phase of Akt signaling.

Small molecule-induced cytosolic activation of protein kinase Akt rescues ischemia-elicited neuronal death.

Elevating Akt activation is an obvious clinical strategy to prevent progressive neuronal death in neurological diseases. However, this endeavor has been hindered because of the lack of specific Akt activators. Here, from a cell-based high-throughput chemical genetic screening, we identified a small molecule SC79 that inhibits Akt membrane translocation, but paradoxically activates Akt in the cytosol.

Exploiting effectors of Rac GTPase.

Targeting a specific arm of signaling pathways is of great interest. In this issue of Chemistry & Biology, Bosco et al. exploit the interactive interface between Rac GTPase and its effector p67(phox) to specifically inhibit reactive oxygen species production without perturbing other Rac-mediated cellular processes.

Deactivation of Akt by a small molecule inhibitor targeting pleckstrin homology domain and facilitating Akt ubiquitination.

The phosphatidylinositol-3,4,5-triphosphate (PIP3) binding function of pleckstrin homology (PH) domain is essential for the activation of oncogenic Akt/PKB kinase. Following the PIP3-mediated activation at the membrane, the activated Akt is subjected to other regulatory events, including ubiquitination-mediated deactivation. Here, by identifying and characterizing an allosteric inhibitor, SC66, we show that the facilitated ubiquitination effectively terminates Akt signaling.

Natural product Celastrol destabilizes tubulin heterodimer and facilitates mitotic cell death triggered by microtubule-targeting anti-cancer drugs.

Background: Microtubule drugs are effective anti-cancer agents, primarily due to their ability to induce mitotic arrest and subsequent cell death. However, some cancer cells are intrinsically resistant or acquire a resistance. Lack of apoptosis following mitotic arrest is thought to contribute to drug resistance that limits the efficacy of the microtubule-targeting anti-cancer drugs.

Small-molecule screen identifies reactive oxygen species as key regulators of neutrophil chemotaxis.

Neutrophil chemotaxis plays an essential role in innate immunity, but the underlying cellular mechanism is still not fully characterized. Here, using a small-molecule functional screening, we identified NADPH oxidase-dependent reactive oxygen species as key regulators of neutrophil chemotactic migration. Neutrophils with pharmacologically inhibited oxidase, or isolated from chronic granulomatous disease (CGD) patients and mice, formed more frequent multiple pseudopodia and lost their directionality as they migrated up a chemoattractant concentration gradient.

Cancer cell-derived clusterin modulates the phosphatidylinositol 3'-kinase-Akt pathway through attenuation of insulin-like growth factor 1 during serum deprivation.

Cancer cells in their respective microenvironments must endure various growth-constraining stresses. Under these conditions, the cancer cell-derived factors are thought to modulate the signaling pathways between cell growth and dormancy. Here, we describe a cancer cell-derived regulatory system that modulates the phosphatidylinositol 3'-kinase (PI3K)-Akt pathway under serum deprivation stress.

Inositol 1,3,4,5-tetrakisphosphate negatively regulates phosphatidylinositol-3,4,5- trisphosphate signaling in neutrophils.

Many neutrophil functions are regulated by phosphatidylinositol-3,4,5-trisphosphate (PtdIns(3,4,5)P3) that mediates protein membrane translocation via binding to pleckstrin homolog (PH) domains within target proteins. Here we show that inositol 1,3,4,5-tetrakisphosphate (Ins(1,3,4,5)P4), a cytosolic small molecule, bound the same PH domain of target proteins and competed for binding to PtdIns(3,4,5)P3. In neutrophils, chemoattractant stimulation triggered rapid elevation in Ins(1,3,4,5)P4 concentration.

RNAi screen identifies UBE2D3 as a mediator of all-trans retinoic acid-induced cell growth arrest in human acute promyelocytic NB4 cells.

All-trans retinoic acid (ATRA) has been widely used in differentiation therapy for acute promyelocytic leukemia (APL). ATRA binds to retinoic acid receptor (RAR) and triggers the formation of the transcription coactivator complex, which leads to changes in gene expression, APL cell-cycle arrest and differentiation, and clinical remission. The mechanisms responsible for ATRA's beneficial effects are still ill-defined.

Tumor suppressor PTEN is a physiologic suppressor of chemoattractant-mediated neutrophil functions.

The recruitment and activation of neutrophils at infected tissues is essential for host defense against invading microorganisms. However, excessive neutrophil recruitment or activation can also damage the surrounding tissues and cause unwanted inflammation. Hence, the responsiveness of neutrophils needs to be tightly regulated.

Deactivation of phosphatidylinositol 3,4,5-trisphosphate/Akt signaling mediates neutrophil spontaneous death.

Neutrophil spontaneous death plays essential roles in neutrophil homeostasis and resolution of inflammation, whereas the underlying molecular mechanisms are still ill-defined. Neutrophils die because of programmed cell death or apoptosis. However, treatment with inhibitor of caspases, which are responsible for the majority of apoptotic cell deaths, does not prevent the spontaneous death of neutrophils.

Facilitated forward chemical genetics using a tagged triazine library and zebrafish embryo screening.

An improved forward chemical genetics approach was successfully demonstrated using a tagged library concept. A small-molecule triazine library with linkers was used to screen for brain/eye developmental phenotypes in a zebrafish embryo system. This approach enabled the rapid isolation of the target proteins by facile affinity matrix preparation and elucidated the first small-molecule inhibitors for several ribosomal accessory proteins or their complex as the target.

Zebrafish N-cadherin, encoded by the glass onion locus, plays an essential role in retinal patterning.

Genetic screens in zebrafish identified several loci that play essential roles in the patterning of retinal architecture. Here, we show that one of them, glass onion, encodes the N-cadherin gene. The glo(m117) mutant allele contains a substitution of the Trp2 residue known for its essential role in the adhesive properties of classic cadherins.

Analysis of gene function in the zebrafish retina.

Mutagenesis screens in zebrafish have uncovered several hundred mutant alleles affecting the development of the retina and established the zebrafish as one of the leading models of vertebrate eye development. In addition to forward genetic mutagenesis approaches, gene function in the zebrafish embryo is being studied using several reverse genetic techniques. Some of these rely on the overexpression of a gene product, others take advantage of antisense oligonucleotides to block function of selected loci.