Soojin Hwang

Drosophila DJ-1 decreases neural sensitivity to stress by negatively regulating Daxx-like protein through dFOXO.

DJ-1, a Parkinsons disease (PD)–associated gene, has been shown to protect against oxidative stress in Drosophila. However, the molecular mechanism underlying oxidative stress-induced phenotypes, including apoptosis, locomotive defects, and lethality, in DJ-1-deficient flies is not fully understood. Here we showed that Daxx-like protein (DLP), a Drosophila homologue of the mammalian Death domain-associated protein (Daxx), was upregulated under oxidative stress conditions in the loss-of-function mutants of Drosophila DJ-1β, a Drosophila homologue of DJ-1. DLP overexpression induced apoptosis via the c-Jun N-terminal kinase (JNK)/Drosophila forkhead box subgroup O (dFOXO) pathway, whereas loss of DLP increased resistance to oxidative stress and UV irradiation. Moreover, the oxidative stress-induced phenotypes of DJ-1β mutants were dramatically rescued by DLP deficiency, suggesting that enhanced expression of DLP contributes to the DJ-1β mutant phenotypes. Interestingly, we found that dFOXO was required for the increase in DLP expression in DJ-1β mutants and that dFOXO activity was increased in the heads of DJ-1β mutants. In addition, subcellular localization of DLP appeared to be influenced by DJ-1 expression so that cytosolic DLP was increased in DJ-1β mutants. Similarly, in mammalian cells, Daxx translocation from the nucleus to the cytosol was suppressed by overexpressed DJ-1β under oxidative stress conditions; and, furthermore, targeted expression of DJ-1β to mitochondria efficiently inhibited the Daxx translocation. Taken together, our findings demonstrate that DJ-1β protects flies against oxidative stress- and UV-induced apoptosis by regulating the subcellular localization and gene expression of DLP, thus implying that Daxx-induced apoptosis is involved in the pathogenesis of DJ-1-associated PD.

Neuroprotective effect of SuHeXiang Wan in Drosophila models of Alzheimer's disease.

AIM OF THE STUDY SuHeXiang Wan (SHXW) is a Chinese traditional medicinal prescription that consists of 15 crude herbs. SHXW has been used to treat central nervous depression, seizures, infantile convulsion and stroke, and its essential oil has been shown to have anticonvulsant and antioxidative activity. The goal of this study was to investigate the beneficial effects of SHXW on the neurological phenotypes of Drosophila AD models. MATERIALS AND METHODS We evaluated the effects of a modified SHXW (called KSOP1009) intake on the AD-like phenotypes of Drosophila AD models, which express human Aβ42 in their developing eyes or neurons. RESULTS When the flies were kept on the media containing 5 μg/ml of KSOP1009 extract, Aβ42-induced eye degeneration, apoptosis, and the locomotive dysfunctions were strongly suppressed. However, Aβ42 fibril deposits in the Aβ42 overexpressing model were not affected by treatment with KSOP1009 extract. Conversely, KSOP1009 extract intake significantly suppressed the constitutive active form of hemipterous, a JNK activator, while it induced eye degeneration and JNK activation, which has been recognized as an important mediator of Aβ42-associated neuro-cytotoxicity. CONCLUSIONS In conclusion, the results of this study suggest that KSOP1009 confers a therapeutic potential to AD-like pathology of Aβ42 overexpressing Drosophila model via suppression of the hyperactivation of JNK activity and apoptosis.

Parkin Suppresses c-Jun N-terminal kinase-induced cell death via transriptional regulation in Drosophila

Parkin is the most prevalent genetic factor in the onset of autosomal recessive juvenile parkinsonism (AR-JP), and mutations in parkin has been reported to cause motor defects, which result from dopamine deficiency caused by dopaminergic neuronal cell death. Activation of c-Jun N-terminal kinase (JNK) has also been implicated in neuronal cell death in Parkinson’s disease (PD). Moreover, Drosophila models for AR-JP, loss of function mutants of Drosophila parkin, also show dopaminergic neural degeneration associated with hyperactivation of JNK, increased apoptosis, and mitochondrial defects. However, the molecular mechanism by which Parkin protects cells from apoptosis remains unclear. In the present study, we tested whether Drosophila Parkin suppressed the JNK signaling pathway in developing tissues. Ectopically expressed parkin strongly suppressed the constitutively active form of Hemipterous (HepCA), a Drosophila JNK kinase that induces an eye degeneration phenotype and apoptosis in the eye imaginal disc. Moreover, parkin also suppressed extra vein formation induced by Basket (Bsk), a Drosophila JNK. Interestingly, the bsk mRNA level was markedly reduced by parkin over-expression, suggesting that the effect of parkin on the phenotype induced by activation of JNK signaling was achieved by transcriptional regulation. Furthermore, we found that the expression level of JNK target genes was reduced by parkin over-expression. Taken together, these results suggest that Drosophila Parkin suppresses JNK signaling by reducing bsk transcription.

The effects of hempseed meal intake and linoleic acid on Drosophila models of neurodegenerative diseases and hypercholesterolemia.

Hempseed is rich in polyunsaturated fatty acids (PUFAs), which have potential as therapeutic compounds for the treatment of neurodegenerative and cardiovascular disease. However, the effect of hempseed meal (HSM) intake on the animal models of these diseases has yet to be elucidated. In this study, we assessed the effects of the intake of HSM and PUFAs on oxidative stress, cytotoxicity and neurological phenotypes, and cholesterol uptake, using Drosophila models. HSM intake was shown to reduce H2O2 toxicity markedly, indicating that HSM exerts a profound antioxidant effect. Meanwhile, intake of HSM, as well as linoleic or linolenic acids (major PUFA components of HSM) was shown to ameliorate Aβ42-induced eye degeneration, thus suggesting that these compounds exert a protective effect against Aβ42 cytotoxicity. On the contrary, locomotion and longevity in the Parkinson’s disease model and eye degeneration in the Huntington’s disease model were unaffected by HSM feeding. Additionally, intake of HSM or linoleic acid was shown to reduce cholesterol uptake significantly. Moreover, linoleic acid intake has been shown to delay pupariation, and cholesterol feeding rescued the linoleic acid-induced larval growth delay, thereby indicating that linoleic acid acts antagonistically with cholesterol during larval growth. In conclusion, our results indicate that HSM and linoleic acid exert inhibitory effects on both Aβ42 cytotoxicity and cholesterol uptake, and are potential candidates for the treatment of Alzheimer’s disease and cardiovascular disease.

Dietary hempseed meal intake increases body growth and shortens the larval stage via the upregulation of cell growth and sterol levels in Drosophila melanogaster

Hempseed, a rich source of polyunsaturated fatty acids (PUFAs) and phytosterols, has been recognized as a potential therapeutic food used for cardioprotection, preventing platelet aggregation, and improving atopic dermatitis. Although several studies have revealed the physiological benefits of hempseed on a variety of animals, the effects of dietary hempseed intake on animal development are currently unknown. In this study, we evaluated the developmental effects of the addition of hempseed meal (HSM) to the diet of Drosophila. Interestingly, dietary HSM intake was shown to increase the body size of flies by increasing cell numbers, and also truncated the larval period without affecting survival rate or longevity. The oviposition of female flies was also increased by dietary HSM supplementation. Interestingly, the levels of sterols, which are precursors of ecdysone, a molting hormone, were found to be elevated in the larvae fed on HSM. Additionally, the hexane extracts of hempseed mimicked the effects of HSM on growth, developmental timing, and reproduction. Moreover, among the major nonpolar components of HSM, feeding on cholesterol but not PUFA mix or campesterol accelerated pupariation and increased body size. These results indicate that the dietary intake of HSM accelerates both body growth and developmental rates in Drosophila via the stimulation of cell growth and ecdysone synthesis. Additionally, nonpolar components of hempseed, such as cholesterol, might be responsible for the effects of HSM on development and reproduction.

dXNP/DATRX increases apoptosis via the JNK and dFOXO pathway in Drosophila neurons.

Mutation of the XNP/ATRX gene, which encodes an SNF2 family ATPase/helicase protein, leads to ATR-X syndrome and several other X-linked mental retardation syndromes. Although XNP/ATRX is a chromatin remodeler, the molecular mechanism by which mental retardation occurs in patients with ATR-X has yet to be determined. To better understand the role of XNP/ATRX in neuronal development, we expressed Drosophila XNP (dXNP/DATRX) ectopically in Drosophila neurons. Neuronal expression of dXNP/DATRX resulted in various developmental defects and induced strong apoptosis. These defects and apoptosis were suppressed by Drosophila inhibitor of apoptosis protein 1. Expression of dXNP/DATRX also increased JNK activity and the levels of reaper and hid transcripts, which are pro-apoptotic factors that activate caspase. Furthermore, dXNP/DATRX-induced rough eye phenotype and apoptosis were suppressed by dFOXO deficiency. These results suggest that dXNP/DATRX is involved in caspase-dependent apoptosis in Drosophila neurons via regulation of the JNK and dFOXO pathway.

Tsp66E, the Drosophila KAI1 homologue, and Tsp74F function to regulate ovarian follicle cell and wing development by stabilizing integrin localization

The metastasis suppressor KAI1/CD82 has been implicated in various cellular processes; however, its function in development is not fully understood. Here, we generated and characterized mutants of Tsp66E and Tsp74F, which are Drosophila homologues of KAI1/CD82 and Tspan11, respectively. These mutants exhibited egg elongation defects along with disturbed integrin localization and actin polarity. Moreover, the defects were enhanced by mutation of inflated, an αPS2 integrin gene. Mutant ovaries had elevated αPS2 integrin levels and reduced endocytic trafficking. These results suggest that Drosophila KAI1/CD82 affects the polarized localization and the level of integrin, which may contribute to epithelial cell polarity.

Effects of sarah/nebula knockdown on Aβ42-induced phenotypes during Drosophila development

The Down syndrome critical region 1 (DSCR1), a Down syndrome-associated protein, is an endogenous inhibitor of the Ca2+-dependent phosphatase calcineurin. It has been also suggested to be associated with Alzheimer’s disease (AD) but the role of DSCR1 in the pathogenesis of AD still remains controversial. In this paper, we investigated the effects of knockdown of sarah (sra), a DrosophilaDSCR1 ortholog, on the Aβ42-induced developmental phenotypes of Drosophila. Knockdown of sra showed detrimental effects on the rough eye phenotype and survival of Aβ42-expressing flies without altering the Aβ42 accumulation. Furthermore, the knockdown of sra increased glial cell numbers in the larval brains and its susceptibility to oxidative stress. Overexpression of an active form of calcineurin produced similar results to sra knockdown as they both exacerbated the Aβ42-induced rough eye phenotype. However, sra knockdown did not alter apoptosis or c-Jun N-terminal kinase activation in Aβ42-expressing flies. In conclusion, our results suggest that sra does play an important role in Aβ42-induced developmental defects in Drosophila without affecting its stress responses.

Interference in xbp1 gene expression induces defective cell differentiation and sensory organ development in Drosophila

The X-box binding protein 1 (Xbp1) is a transcription factor that is important in the unfolded protein response to protect cells from endoplasmic reticulum (ER) stress and is implicated in several other biological processes, including liver growth and B lymphocyte differentiation. Although the cellular function of Xbp1 has been well studied, its developmental role remains to be elucidated. In the present study, we investigated the developmental role of Drosophila xbp1 using an RNAi-mediated gene silencing system. When xbp1 gene expression was specifically interfered with in developing tissues, such as wing and eye during larval growth, the tissues were strongly deformed. Cell death was not observed in these tissues, and this phenotype was not rescued by the overexpression of Drosophila inhibitor of apoptosis protein1 (DIAP1), a caspase inhibitor, indicating that these developmental defects are not associated with apoptosis. Furthermore, changes were not observed in cell cycle progression and Jun-N-terminal kinase activation in tissues whose xbp1 gene expression was interfered with. Rather, the xbp1 gene silencing resulted in defective cell differentiation and supernumerary of sensory organ precursors. These results suggest the inability of xbp1 gene silencing to induce strong ER stress and the importance of Drosophila Xbp1 in cell fate determination and sensory organ development.

Phenotypic differences between Drosophila Alzheimer’s disease models expressing human Aβ42 in the developing eye and brain

ABSTRACT Drosophila melanogaster expressing amyloid-β42 (Aβ42) transgenes have been used as models to study Alzheimer’s disease. Various Aβ42 transgenes with different structures induce different phenotypes, which make it difficult to compare data among studies which use different transgenic lines. In this study, we compared the phenotypes of four frequently used Aβ42 transgenic lines, UAS-Aβ422X, UAS-Aβ42BL33770, UAS-Aβ4211C39, and UAS-Aβ42H29.3. Among the four transgenic lines, only UAS-Aβ422X has two copies of the upstream activation sequence-amyloid-β42 (UAS-Aβ42) transgene, while remaining three have one copy. UAS-Aβ42BL33770 has the 3′ untranslated region of Drosophila α-tubulin, while the others have that of SV40. UAS-Aβ4211C39 and UAS-Aβ42H29.3 have the rat pre-proenkephalin signal peptide, while UAS-Aβ422X and UAS-Aβ42BL33770 have that of the fly argos protein. When the transgenes were expressed ectopically in the developing eyes of the flies, UAS-Aβ422X transgene resulted in a strongly reduced and rough eye phenotype, while UAS-Aβ42BL33770 only showed a strong rough eye phenotype; UAS-Aβ42H29.3 and UAS-Aβ4211C39 had mild rough eyes. The levels of cell death and reactive oxygen species (ROS) in the eye imaginal discs were consistently the highest in UAS-Aβ422X, followed by UAS-Aβ42BL33770, UAS-Aβ4211C39, and UAS-Aβ42H29.3. Surprisingly, the reduction in survival during the development of these lines did not correlate with cell death or ROS levels. The flies which expressed UAS-Aβ4211C39 or UAS-Aβ42H29.3 experienced greatly reduced survival rates, although low levels of ROS or cell death were detected. Collectively, our results demonstrated that different Drosophila AD models show different phenotypic severity, and suggested that different transgenes may have different modes of cytotoxicity. Abbreviations: Aβ42: amyloid-β42; AD: Alzheimer’s disease; UAS: upstream activation sequence