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Park, Seo, and Chun: Targeted Downregulation of kdm4a Ameliorates Tau-engendered Defects in Drosophila melanogaster
Original Article
Basic Medical Sciences

Journal of Korean Medical Science 2019; 34(33): e225.

Published online: 8 August 2019

DOI: https://doi.org/10.3346/jkms.2019.34.e225

Targeted Downregulation of kdm4a Ameliorates Tau-engendered Defects in Drosophila melanogaster

Sung Yeon Park1, 2, Jieun Seo2, 3, Yang-Sook Chun1, 2, 3

1Ischemic/Hypoxic Disease Institute, Seoul National University College of Medicine, Seoul, Korea.

2Department of Physiology, Seoul National University College of Medicine, Seoul, Korea.

3Department of Biomedical Sciences, Seoul National University College of Medicine, Seoul, Korea.

Address for Correspondence: Yang-Sook Chun, PhD. Ischemic/Hypoxic Disease Institute, Department of Physiology and Department of Biomedical Sciences, Seoul National University College of Medicine, 103 Daehak-ro, Jongno-gu, Seoul 03080, Korea. chunys@snu.ac.kr

Received 17 June 2019       Accepted 22 July 2019

© 2019 The Korean Academy of Medical Sciences.

The Korean Academy of Medical Sciences

(open-access):

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (https://creativecommons.org/licenses/by-nc/4.0/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Background

Tauopathies, a class of neurodegenerative diseases that includes Alzheimer's disease (AD), are characterized by the deposition of neurofibrillary tangles composed of hyperphosphorylated tau protein in the human brain. As abnormal alterations in histone acetylation and methylation show a cause and effect relationship with AD, we investigated the role of several Jumonji domain-containing histone demethylase (JHDM) genes, which have yet to be studied in AD pathology.

Methods

To examine alterations of several JHDM genes in AD pathology, we performed bioinformatics analyses of JHDM gene expression profiles in brain tissue samples from deceased AD patients. Furthermore, to investigate the possible relationship between alterations in JHDM gene expression profiles and AD pathology in vivo, we examined whether tissue-specific downregulation of JHDM Drosophila homologs (kdm) can affect tauR406W-induced neurotoxicity using transgenic flies containing the UAS-Gal4 binary system.

Results

The expression levels of JHDM1A, JHDM2A/2B, and JHDM3A/3B were significantly higher in postmortem brain tissue from patients with AD than from non-demented controls, whereas JHDM1B mRNA levels were downregulated in the brains of patients with AD. Using transgenic flies, we revealed that knockdown of kdm2 (homolog to human JHDM1), kdm3 (homolog to human JHDM2), kdm4a (homolog to human JHDM3A), or kdm4b (homolog to human JHDM3B) genes in the eye ameliorated the tauR406W-engendered defects, resulting in less severe phenotypes. However, kdm4a knockdown in the central nervous system uniquely ameliorated tauR406W-induced locomotion defects by restoring heterochromatin.

Conclusion

Our results suggest that downregulation of kdm4a expression may be a potential therapeutic target in AD.

Graphical Abstract

jkms-34-e225-abf001

Keywords: Drosophila melanogaster, Tauopathy, Alzheimer's Disease, Heterochromatin, JHDM, kdm

INTRODUCTION

Tauopathies, a class of neurodegenerative diseases that includes Alzheimer's disease (AD), are characterized by the deposition of neurofibrillary tangles composed of hyperphosphorylated tau protein in the human brain.1 However, the underlying mechanism of tauopathy remains unclear, and deciphering the process is crucial for developing effective biomarkers and identifying potential therapeutic targets of AD. Abnormal alterations of histone acetylation and methylation have been documented in patients with AD and in a transgenic animal model of AD.2 In early development, several epigenetic alterations may contribute to AD, whereas certain epigenetic shifts may occur downstream of AD pathology.2 In particular, histone acetylation is indicative of late onset cognitive loss, and has been used to develop biomarkers of AD.3
Tau phosphorylation is an important pathogenic event in AD. Targeted expression of human tau in the central nervous system (CNS) of Drosophila recapitulates several characteristics in human AD, such as neuronal loss in the form of prominent vacuoles, compromised life span, and locomotor and cognitive impairments.4 Therefore, to provide a useful AD model, transgenic Drosophila were generated using an FTDP-17 mutant form of human tau (tauR406W) that was subcloned downstream of the yeast upstream activator sequence (UAS),5 tissue-specific expression of the transgene was achieved by regulating the availability of Gal4 transcription factor with different Gal4 drivers.6
Analyses of tissues from AD patients and a transgenic mouse model of AD consistently show decreased histone acetylation.7 Accordingly, histone deacetylase (HDAC) inhibitors have been proposed as potential procognitive agents for the treatment of AD.891011 Indeed, Tip60 histone acetyltransferase (HAT) in human tau overexpressing Drosophila has been proposed to play a neuroprotective role in impaired cognition during early development of AD.121314 Reciprocally, Drosophila genetic screens have shown that HDAC6 null mutation and pharmacological inhibition can rescue tau-induced microtubule defects in muscles and neurons.15 Furthermore, histone methylation has a marked influence on synaptic plasticity and cognition.81016
Recently, Mastroeni and colleagues reported increased histone H3K4 trimethylation in the cytoplasm and significant colocalization with hyperphosphorylated tau tangles, but decreased in the nuclei of AD brains compared to non-demented controls.17 Moreover, the total levels of histone H3K9 dimethylation and HP1α were decreased and heterochromatin relaxed, resulting in aberrant neuronal gene expression in Drosophila, mouse, and human tauopathies.5 However, no studies have investigated the involvement of histone demethylases in tauopathies.
In this study, bioinformatics analyses revealed significantly higher expression of several Jumonji domain-containing histone demethylase (JHDM) genes18192021 in postmortem brain tissue from patients with AD than from non-demented controls. In addition, our results suggest that the downregulation of kdm4a (JHDM3A) expression may be a potential therapeutic target in AD, providing new insight into the role of histone demethylases in tauopathies.

METHODS

Bioinformatics analyses

Expression levels of histone demethylases in postmortem brain tissue from patients with AD were evaluated using the publically available Gene Expression Omnibus (www.ncbi.nlm.nih.gov/geo, GSE33000) (Table 1). The dataset included patients with AD (n = 310) and non-demented controls (n = 157). JHDM1A (10025907153_at probe), JHDM1B (10025912776_at probe), JHDM2A (10023810558_at probe), JHDM2B (10025911102_at probe), JHDM3A (10025904057_at probe) and JHDM3B (10025924555_at probe) mRNA expression levels were compared between groups using unpaired, two-sided Student's t-tests.
Table 1

GSE33000 subject information

jkms-34-e225-i001
Classification Control subjects AD subjects
Tissue source Prefrontal cortex Prefrontal cortex
No. 157 310
Age, yr 63.0 ± 10.2 80.0 ± 9.8
Sex, male/female 123/34 135/175
AD = Alzheimer's disease.

Drosophila strains and genetics

Fruit flies were maintained on standard media, and genetic crosses were performed at 25°C unless specified otherwise. All genetic crosses were performed at least in triplicate, and percentages represent the means of the replicates. The following Drosophila strains were used: UAS-kdm2 RNAi (P{KK101783}VIE-260B) (Vienna Drosophila RNAi Center [VDRC], Vienna, Austria), UAS-kdm3 RNAi (y[1] v[1]; P{y[+t7.7] v[+t1.8] =TRiP.HMJ22328}attP40) (Bloomington Drosophila Stock Center, Indiana University, Bloomington, IN, USA), UAS-kdm4a RNAi lines (w1118; P{GD9133}v32652 hereafter referred to as UAS-kdm4a RNAi and w1118; P{GD9133}v32650/CyO hereafter referred to as UAS-kdm4a RNAi2) (VDRC), UAS-kdm4b RNAi (P{KK102089}VIE-260B) (VDRC), elav-Gal4 lines (P{w[+mW.hs]=GawB}elav[C155] and P{w[+mC]=GAL4-elav.L}2/CyO, #B8765) (Bloomington Drosophila Stock Center), UAS-kdm4a-HA1FLAG2 (w; P{w[+mC]=[UAS-kdm4a-HA1FLAG2]} hereafter referred to as UAS-kdm4a+) (Jerry L. Workman, Stowers Institute for Medical Research, Kansas, MO, USA),22 BL2 chromatin reporter (y1·w*/Dp(3;Y)BL2, P{HS-lacZ.scs}65E) (Bloomington Drosophila Stock Center), UAS-tauR406W (Mel B. Feany, Harvard Medical School, Boston, MA, USA),5 GMR-Gal4 (Deborah A. Hursh, CBER/FDA, Silver Springs, MD, USA).23 The wild-type strain was w1118 (Seungbok Lee, Seoul National University, Seoul, Korea).24

Locomotor behavioral assay

Flies were collected in vials on the day of eclosion and then transferred without anesthesia to fresh vials every other day. The locomotor assay was performed in the afternoon on day 10. Ten flies were placed in each plastic vial marked with 15 cm height and gently tapped to the bottom. The percentage of flies that climbed to the height of 15 cm in 10 seconds at each consecutive 5-day interval was calculated for each genotype unless specified otherwise. The assay was performed for 20 days after eclosion.

Histochemical detection

For histochemical detection of β-galactosidase activity, larvae were heat-shocked for 45 minutes at 37°C, followed by recovery for 1 hour at room temperature. Larval brain lobes were dissected in phosphate-buffered saline (PBS), fixed in 3.7% formaldehyde in PBS for 5 minutes, washed in PBS, transferred to X-gal solution,25 and incubated overnight at 37°C. Larval brain lobes were mounted in 80% glycerol and examined using an Olympus DP71 microscope.

Statistical analyses

All data were analyzed using Microsoft Excel 2007 and expressed as the means and standard deviations. Continuous variables were analyzed using Student's t-tests when data were normally distributed. All statistical tests were two-sided. P values < 0.05 were considered to indicate statistical significance.

RESULTS

Increased expression of several JHDM genes in postmortem brain tissue from patients with AD

To investigate the potential relationship between alterations in the expression of JHDM genes and AD pathology in vivo, we used bioinformatics analyses to examine differences in the expression levels of JHDM1A/1B, JHDM2A/2B, and JHDM3A/3B, which have Drosophila homologs (Tables 1 and 2). The expression levels of JHDM1A, JHDM2A/2B, and JHDM3A/3B were significantly higher in postmortem brain tissue from AD patients than from non-demented controls (average log2 value, 0.0221 ± 0.00337 vs. −0.0333 ± 0.00451; 0.0064 ± 0.00516 vs. −0.0794 ± 0.00543; 0.00837 ± 0.00276 vs. −0.0179 ± 0.00342; 0.0471 ± 0.00251 vs. 0.0256 ± 0.00326; 0.1403 ± 0.0155 vs. −0.0220 ± 0.0171, respectively; P < 0.001), whereas the JHDM1B mRNA levels were significantly downregulated in brain tissues from AD patients (average log2 value, −0.1300 ± 0.00603 vs. −0.0161 ± 0.00826; P < 0.001) (Fig. 1).
Table 2

List of genes for the study

jkms-34-e225-i002
Fly symbol Fly gene ID Human symbol Human gene ID DIOPTa score Weighted score Rank
kdm2 cg11033 JHDM1A/KDM2A 22992 14 of 15 13.85 High
JHDM1B/KDM2B 84678 11 of 15 10.97 Moderate
kdm3 cg8165 JHDM2A/KDM3A 55818 9 of 15 8.75 Moderate
JHDM2B/KDM3B 51780 10 of 15 9.76 High
kdm4a cg15835 JHDM3A/KDM4A 9682 10 of 15 9.74 High
JHDM3B/KDM4B 23030 10 of 15 9.79 High
kdm4b cg33182 JHDM3A/KDM4A 9682 10 of 15 9.74 High
JHDM3B/KDM4B 23030 9 of 15 8.79 Moderate
aDrosophila RNAi Screening Center Integrative Orthologue Prediction Tool (ver 7.1).
Fig. 1

Increased expression of JHDM genes in postmortem brain tissue from patients with AD. Scatter diagrams representing mRNA expression levels of JHDM genes in postmortem brain tissue from patients with AD. Dot plots showing JHDM1A/1B, JHDM2A/2B, and JHDM3A/3B mRNA levels in non-demented healthy controls and AD patients.

JHDM = Jumonji domain-containing histone demethylase, AD = Alzheimer's disease.
*P < 0.001 between the indicated groups.
jkms-34-e225-g001

Knockdown of JHDM genes ameliorates the morphological deficit induced by overexpressing tauR406W in Drosophila eyes

To examine the effect of JHDM gene knockdown in an AD background, tauR406W-expressing flies were crossed with UAS-RNAi strains expressing RNAi specific for each of the JHDM fly homologs (kdm2, JHDM1; kdm3, JHDM2; kdm4a, JHDM3A; and kdm4b, JHDM3B) (Table 2) in Drosophila eye using the GMR-Gal4 driver. Neurodegeneration can be easily monitored when tauR406W is overexpressed in the fly eyes, resulting in eye morphological defects such as reduced size, loss of bristles, disordered ommatidia and roughened eye surfaces.2627 The observed eye defects can be categorized into four classes according to their severity: class 1, normal; class 2, roughness in the eyes with size reduction and some ommatidial disruption; class 3, collapsed eye tissue with size reduction; and class 4, black spots of apoptotic tissue in the collapsed eye (Fig. 2A). Approximately 74% of flies overexpressing tauR406W have class 3 + 4 eyes. In contrast, knockdown of kdm2, kdm3, kdm4a or kdm4b genes in tauR406W-overexpressing flies ameliorated the tauR406W-engendered eye defects, resulting in only 8%, 37%, 40%, or 21% class 3 + 4 eyes, respectively (Fig. 2B). No eye defects were detected in flies overexpressing each of the UAS-RNAi strains of kdm alone (data not shown). Therefore, knockdown of the kdm genes ameliorated the tauR406W-induced eye phenotype, resulting in less severe phenotypes.
Fig. 2

Knockdown of any of the JHDM genes ameliorates the morphological defects induced by overexpressing tauR406W in Drosophila eyes. (A) Bright field images of 10-day-old adult Drosophila eyes of control (w1118) and tauR406W transgenic flies heterozygous for knockdown of any of the kdm genes using the GMR-Gal4. Images were taken using an Olympus SZ61 stereo zoom binocular microscope equipped with an eXcope XCAM1080 digital camera. (B) kdm knockdown flies that overexpressed tauR406W showed a shift toward less severity in the distribution of phenotype categories in a population of flies (n = 30–73).

JHDM = Jumonji domain-containing histone demethylase.
jkms-34-e225-g002

Targeted downregulation of kdm4a ameliorates tauR406W-induced locomotion defect

To evaluate the neuronal effect of kdm gene knockdown in flies overexpressing tauR406W in their CNS via pan-neuronal elav-Gal4, we performed the climbing assay, a behavioral readout used for monitoring neurotoxic deficits in Drosophila.2829303132 Whereas w1118 control flies retained 90% of their climbing ability on day 10, only 45%–50% of tauR406W-expressing flies climbed up. Inconsistent with the eye analysis results, knockdown of either kdm2 or kdm3 in flies overexpressing tauR406W had no significant effect on the climbing ability of flies overexpressing tauR406W alone (Fig. 3A and B). Interestingly, flies that overexpressed kdm2 RNAi alone exhibited a locomotion defect, showing 60% climbing ability compared to 90% climbing ability in control flies. This finding indicates that kdm2 itself may play a role in neuronal function. However, knockdown of kdm4a in tauR406W-overexpressing flies caused a mild but significant amelioration of the locomotion defect, with flies showing 80% climbing ability compared to the 50%–60% climbing ability of flies overexpressing tauR406W alone (Fig. 3C). Importantly, overexpression of kdm4a RNAi alone had no significant effect on locomotion, indicating that the neuroprotective effect may be specific to the context of tauopathy. In contrast, knockdown of kdm4b in flies that overexpressed tauR406W exacerbated the locomotion defect, and they showed 30% climbing ability compared to 50% in flies overexpressing tauR406W alone. Moreover, flies expressing kdm4b RNAi alone exhibited the locomotion defect, showing 60% climbing ability compared to 90% in control flies, indicating that kdm4b itself may be involved in neuronal function (Fig. 3D). Therefore, tauR406W overexpression may aggravate the neuronal defect in kdm4b-deficient flies during development.
Fig. 3

Targeted downregulation of kdm4a ameliorates tauR406W-induced locomotion defect. (A–E) Locomotor activity in control and tauR406W transgenic flies heterozygous for knockdown of any of the kdm genes using pan-neuronal elav-Gal4 (#8765) was measured on day 10 post-eclosion (10 flies and 10 repeats per group, respectively; 15 cm/10 sec). (F) Locomotor activity in control and tauR406W transgenic flies heterozygous for overexpression of wild-type kdm4a transgene using pan-neuronal elav-Gal4 (#c155) was measured on day 7 post-eclosion (10 flies and 10 repeats per group, respectively; 8 cm/20 sec). (G) A shift in the distribution of eye phenotype categories in populations of flies with kdm4a knockdown or overexpression in tauR406W-overexpressing flies using GMR-Gal4 (n = 78–89).

*P < 0.05 and n.s. denotes P > 0.05 between the indicated groups.
jkms-34-e225-g003
To ascertain the neuronal protective effect of kdm4a knockdown in tauR406W-overexpressing flies, we performed the climbing assay with different kdm4a RNAi strain or wild-type kdm4a transgenic flies crossed to tauR406W-overexpressing flies. Crossing the kdm4a RNAi2 line with flies overexpressing tauR406W in their CNS ameliorated the locomotion defect, as they showed 70% climbing ability compared to 50% in flies that overexpressed tauR406W alone (Fig. 3E), whereas exogenously overexpressing wild-type kdm4a in tauR406W-overexpressing flies exacerbated the locomotion defect, as they showed 10% climbing ability compared to 40% in flies overexpressing tauR406W alone (Fig. 3F). Accordingly, in the eye, kdm4a knockdown with the RNAi2 strain in tauR406W-overexpressing flies resulted in 58.5% class 3 + 4 eyes compared to 87% class 3 + 4 eyes in flies overexpressing tauR406W alone, whereas overexpressing kdm4a in tauR406W-overexpressing flies showed no significant difference in eye morphology compared to those overexpressing tauR406W alone (Fig. 3G). Taken together, kdm4a knockdown ameliorated tauR406W-engendered deficits in both neurons and eyes.

Targeted downregulation of kdm4a reduces heterochromatic loss induced by tauR406W overexpression

Given that tau promotes neuronal death through heterochromatin loss, global chromatin relaxation, and aberrant transcriptional activation in Drosophila, mouse, and human tauopathies,5 we investigated whether knockdown of kdm genes alters heterochromatin loss in flies overexpressing tauR406W in their CNS via pan-neuronal elav-Gal4. We measured reporter expression in larval brain lobes using a transgenic fly strain (BL2 reporter) that has a LacZ reporter gene embedded in and silenced by heterochromatin.533 After 45 min heat shock pulse at 37°C, this LacZ reporter was silenced in controls (Fig. 4A), but abnormally expressed in tauR406W-overexpressing fly brain due to heterochromatin loosening (Fig. 4B). kdm2 knockdown enhanced the expression of the heterochromatin-embedded reporter compared to overexpressed tauR406W alone (Fig. 4C and F). Expectedly, RNAi-mediated reduction of kdm4a decreased the number and intensity of the expression foci of the heterochromatin-embedded reporter in tauR406W-overexpressing flies (Fig. 4D and F), whereas kdm4b knockdown had no significant alteration on the expression of the heterochromatin-embedded reporter in flies overexpressing tauR406W alone (Fig. 4E). Therefore, neuronal-specific downregulation of kdm4a may suppress tauR406W-engendered locomotion impairment by restoring heterochromatin.
Fig. 4

Targeted downregulation of kdm4a reduces heterochromatin loss induced by tauR406Woverexpression. (A–E) The lacZ expression (BL2 reporter) embedded in and silenced by heterochromatin displayed in larval brain lobes of each of the indicated genotypes driven by pan-neuronal elav-Gal4 (#8765) under heat shock pulse (45 min at 37°C), using X-gal staining. (F) Quantification of BL2 chromatin reporter expression of indicated genotypes (n = 5–10).

*P < 0.05 and n.s. denotes P > 0.05 between the indicated groups. Scale bar = 100 μM.
jkms-34-e225-g004

DISCUSSION

We used bioinformatics analyses to detect alterations in JHDM gene expression in AD pathology. The expression levels of JHDM1A, JHDM2A/2B, and JHDM3A/3B were significantly higher in postmortem brain tissue from AD patients than from non-demented controls. In Drosophila, CNS-specific knockdown of kdm4a (JHDM3A) ameliorated tauR406W-induced locomotion defects, in agreement with eye analysis results.
Recently, Frost et al.5 reported that oxidative stress and subsequent DNA damage can substantially alter chromatin structure, inducing heterochromatin loss and aberrant transcriptional activity, causing neurodegeneration in tauopathies. The authors reported that tauR406W-overexpressing adult fly brain had decreased total levels of H3K9 dimethylation and heterochromatin protein 1α (HP1α), which are normally enriched in heterochromatin. In addition, loss of function mutations in HP1α or Su(var)3-9, which encodes a histone methyltransferase responsible for H3K9 dimethylation, exacerbated tauR406W-engendered locomotion deficits. Given that KDM4A also directly interacts with HP1α and is essential for heterochromatin organization and function via both enzymatic and structural mechanisms in Drosophila,223435 it is plausible that KDM4A may be associated with heterochromatin alteration in tauopathies. Intriguingly, kdm4a knockdown with either of two RNAi strains ameliorated tauR406W-induced deficits in the eye and in neurons. Conversely, wild-type kdm4a overexpression in flies overexpressing tauR406W in the CNS exacerbated tauR406W-induced locomotion defects. Importantly, overexpression of kdm4a RNAi alone had no significant effect on locomotion in flies, indicating that the neuroprotective effect was specific for the AD pathological context. Consistent with these results, Lorbeck et al.36 reported that flies containing the P-element suppressor mutant of kdm4a (kdm4aP-supp) showed no significant loss in climbing ability. According to the BL2 transgenic reporter assay, RNAi-mediated reduction of kdm4a in the CNS reduced heterochromatin relaxation, thus ameliorating the tauR406W-engendered locomotion defect. Further studies are needed to elucidate the underlying mechanism of KDM4A involvement in the heterochromatic organization and transcriptional regulation of tauopathies.
kdm4b knockdown in the CNS exacerbated the tauR406W-induced locomotion defect. Interestingly, flies that overexpressed kdm4b RNAi alone exhibited the locomotion defect, showing 60% climbing ability compared to 90% in control flies. This finding indicates that KDM4B may play a role in neuronal function in Drosophila. Consistent with this result, neuronal-specific kdm4b-deficient mice exhibit hyperactive behavior, sustained hyperactivity in a novel environment, deficits in working memory, and spontaneous epileptic-like seizures, indicating impaired neurodevelopment.37 This finding suggests that KDM4B plays a crucial role in the formation of functional neuronal networks. Therefore, neuronal defects in kdm4b-deficient flies during development may exacerbate tauR406W-engendered locomotion impairment. However, the underlying mechanism does not appear to involve altered heterochromatin structure, as kdm4b knockdown had no significant effect on heterochromatin relaxation induced by tauR406W overexpression.
Inconsistent with the eye analysis results, knockdown of either kdm2 or kdm3 in tauR406W-overexpressing flies had no significant effect on climbing ability in flies that overexpressed tauR406W alone. However kdm2 knockdown tended to slightly exacerbate the tauR406W-induced locomotion defect, particularly in young flies (2–3 days after eclosion), but not significantly in older flies (data not shown). Furthermore, flies expressing kdm2 RNAi alone exhibited the locomotion defect, showing 60% climbing ability compared to 90% in control flies, indicating that KDM2 may play a role in neuronal function. In agreement with this result, kdm2 null mutant adult flies displayed defects in circadian locomotor behavior.38 Thus, a neuronal defect in kdm2-deficient flies during development may exacerbate the tauR406W-engendered locomotion impairment in young flies. Furthermore, kdm2 knockdown in the larval brain lobe can enhance the number and intensity of expression foci of the heterochromatin-embedded reporter in tauR406W-overexpressing flies. These results indicate that neuronal-specific downregulation of kdm2 may exacerbate tauR406W-induced locomotion impairment by enhancing heterochromatin loss during early development; however, the underlying mechanism remains to be elucidated.
Using bioinformatics analyses, we revealed significantly increased expression of JHDM1A, JHDM2A/2B, and JHDM3A/3B in postmortem brain tissue from patients with AD compared to non-demented controls, whereas JHDM1B mRNA levels were downregulated in the brains of AD patients. Using tauR406W-induced transgenic flies as an AD model, we showed that kdm4a knockdown in the CNS ameliorated tauR406W-induced locomotion defects by restoring heterochromatin. These results suggest that the downregulation of kdm4a expression could be a potential therapeutic target in AD.

ACKNOWLEDGMENTS

We thank Vincenzo Pirrotta, Deborah Hursh, Brian Stultz, Jerry L. Workman, Mel B Feany, Seungbok Lee, the Bloomington Stock Center, and the Vienna RNAi stock for supplying stocks. Vincenzo Pirrotta made valuable comments that greatly improved the manuscript. Jieun Seo received a scholarship from the BK21-plus education program of the National Research Foundation of Korea.

Notes

Funding This work was supported by a Korean Health Technology R&D grant (HI15C2695) and National Research Foundation grants from the Korean government (2016R1A2B4013377, 2016R1D1A1A02937353, 2018R1D1A1B07042756, 2018R1A2B6007241, 2018R1A5A2025964).

Disclosure The authors have no potential conflicts of interest to disclose.

Author Contributions

  • Conceptualization: Chun YS, Park SY.

  • Data curation: Park SY, Seo J.

  • Formal analysis: Seo J.

  • Funding acquisition: Chun YS, Park SY.

  • Investigation: Chun YS.

  • Methodology: Park SY, Seo J.

  • Supervision: Chun YS.

  • Writing - original draft: Park SY.

  • Writing - review & editing: Chun YS.

References

1. Lee VM, Goedert M, Trojanowski JQ. Neurodegenerative tauopathies. Annu Rev Neurosci. 2001; 24(1):1121–1159.
crossref
2. Millan MJ. The epigenetic dimension of Alzheimer's disease: causal, consequence, or curiosity? Dialogues Clin Neurosci. 2014; 16(3):373–393.
crossref
3. Zhang K, Schrag M, Crofton A, Trivedi R, Vinters H, Kirsch W. Targeted proteomics for quantification of histone acetylation in Alzheimer's disease. Proteomics. 2012; 12(8):1261–1268.
crossref
4. Chanu SI, Sarkar S. Targeted downregulation of dMyc suppresses pathogenesis of human neuronal tauopathies in drosophila by limiting heterochromatin relaxation and tau hyperphosphorylation. Mol Neurobiol. 2017; 54(4):2706–2719.
crossref
5. Frost B, Hemberg M, Lewis J, Feany MB. Tau promotes neurodegeneration through global chromatin relaxation. Nat Neurosci. 2014; 17(3):357–366.
crossref
6. Osterwalder T, Yoon KS, White BH, Keshishian H. A conditional tissue-specific transgene expression system using inducible GAL4. Proc Natl Acad Sci U S A. 2001; 98(22):12596–12601.
crossref
7. Narayan P, Dragunow M. Alzheimer's disease and histone code alterations. Adv Exp Med Biol. 2017; 978:321–336.
crossref
8. Millan MJ. An epigenetic framework for neurodevelopmental disorders: from pathogenesis to potential therapy. Neuropharmacology. 2013; 68:2–82.
crossref
9. Gräff J, Rei D, Guan JS, Wang WY, Seo J, Hennig KM, et al. An epigenetic blockade of cognitive functions in the neurodegenerating brain. Nature. 2012; 483(7388):222–226.
crossref
10. Day JJ, Sweatt JD. Epigenetic mechanisms in cognition. Neuron. 2011; 70(5):813–829.
crossref
11. Fischer A, Sananbenesi F, Wang X, Dobbin M, Tsai LH. Recovery of learning and memory is associated with chromatin remodelling. Nature. 2007; 447(7141):178–182.
crossref
12. Johnson AA, Sarthi J, Pirooznia SK, Reube W, Elefant F. Increasing Tip60 HAT levels rescues axonal transport defects and associated behavioral phenotypes in a Drosophila Alzheimer's disease model. J Neurosci. 2013; 33(17):7535–7547.
crossref
13. Xu S, Wilf R, Menon T, Panikker P, Sarthi J, Elefant F. Epigenetic control of learning and memory in Drosophila by Tip60 HAT action. Genetics. 2014; 198(4):1571–1586.
crossref
14. Xu S, Elefant F. Tip off the HAT-epigenetic control of learning and memory by Drosophila Tip60. Fly (Austin). 2015; 9(1):22–28.
15. Xiong Y, Zhao K, Wu J, Xu Z, Jin S, Zhang YQ. HDAC6 mutations rescue human tau-induced microtubule defects in Drosophila. Proc Natl Acad Sci U S A. 2013; 110(12):4604–4609.
crossref
16. Jarome TJ, Lubin FD. Histone lysine methylation: critical regulator of memory and behavior. Rev Neurosci. 2013; 24(4):375–387.
crossref
17. Mastroeni D, Delvaux E, Nolz J, Tan Y, Grover A, Oddo S, et al. Aberrant intracellular localization of H3k4me3 demonstrates an early epigenetic phenomenon in Alzheimer's disease. Neurobiol Aging. 2015; 36(12):3121–3129.
18. Park SY, Park JW, Chun YS. Jumonji histone demethylases as emerging therapeutic targets. Pharmacol Res. 2016; 105:146–151.
crossref
19. Zheng Y, Hsu FN, Xu W, Xie XJ, Ren X, Gao X, et al. A developmental genetic analysis of the lysine demethylase KDM2 mutations in Drosophila melanogaster. Mech Dev. 2014; 133:36–53.
crossref
20. Herz HM, Morgan M, Gao X, Jackson J, Rickels R, Swanson SK, et al. Histone H3 lysine-to-methionine mutants as a paradigm to study chromatin signaling. Science. 2014; 345(6200):1065–1070.
crossref
21. Lloret-Llinares M, Carré C, Vaquero A, de Olano N, Azorín F. Characterization of Drosophila melanogaster JmjC+N histone demethylases. Nucleic Acids Res. 2008; 36(9):2852–2863.
crossref
22. Lin CH, Li B, Swanson S, Zhang Y, Florens L, Washburn MP, et al. Heterochromatin protein 1a stimulates histone H3 lysine 36 demethylation by the Drosophila KDM4A demethylase. Mol Cell. 2008; 32(5):696–706.
crossref
23. Song Z, Guan B, Bergman A, Nicholson DW, Thornberry NA, Peterson EP, et al. Biochemical and genetic interactions between Drosophila caspases and the proapoptotic genes rpr, hid, and grim. Mol Cell Biol. 2000; 20(8):2907–2914.
24. Nahm M, Kim S, Paik SK, Lee M, Lee S, Lee ZH, et al. dCIP4 (Drosophila Cdc42-interacting protein 4) restrains synaptic growth by inhibiting the secretion of the retrograde Glass bottom boat signal. J Neurosci. 2010; 30(24):8138–8150.
crossref
25. Blackman RK, Sanicola M, Raftery LA, Gillevet T, Gelbart WM. An extensive 3′ cis-regulatory region directs the imaginal disk expression of decapentaplegic, a member of the TGF-beta family in Drosophila. Development. 1991; 111(3):657–666.
crossref
26. Chatterjee S, Sang TK, Lawless GM, Jackson GR. Dissociation of tau toxicity and phosphorylation: role of GSK-3beta, MARK and Cdk5 in a Drosophila model. Hum Mol Genet. 2009; 18(1):164–177.
27. Sang TK, Jackson GR. Drosophila models of neurodegenerative disease. NeuroRx. 2005; 2(3):438–446.
crossref
28. Chan HY, Bonini NM. Drosophila models of human neurodegenerative disease. Cell Death Differ. 2000; 7(11):1075–1080.
crossref
29. Crowther DC, Kinghorn KJ, Page R, Lomas DA. Therapeutic targets from a Drosophila model of Alzheimer's disease. Curr Opin Pharmacol. 2004; 4(5):513–516.
crossref
30. Scherzer-Attali R, Pellarin R, Convertino M, Frydman-Marom A, Egoz-Matia N, Peled S, et al. Complete phenotypic recovery of an Alzheimer's disease model by a quinone-tryptophan hybrid aggregation inhibitor. PLoS One. 2010; 5(6):e11101.
crossref
31. Crowther DC, Kinghorn KJ, Miranda E, Page R, Curry JA, Duthie FA, et al. Intraneuronal Abeta, non-amyloid aggregates and neurodegeneration in a Drosophila model of Alzheimer's disease. Neuroscience. 2005; 132(1):123–135.
32. Frydman-Marom A, Levin A, Farfara D, Benromano T, Scherzer-Attali R, Peled S, et al. Orally administrated cinnamon extract reduces β-amyloid oligomerization and corrects cognitive impairment in Alzheimer's disease animal models. PLoS One. 2011; 6(1):e16564.
crossref
33. Lu BY, Ma J, Eissenberg JC. Developmental regulation of heterochromatin-mediated gene silencing in Drosophila. Development. 1998; 125(12):2223–2234.
crossref
34. Lin CH, Paulson A, Abmayr SM, Workman JL. HP1a targets the Drosophila KDM4A demethylase to a subset of heterochromatic genes to regulate H3K36me3 levels. PLoS One. 2012; 7(6):e39758.
crossref
35. Colmenares SU, Swenson JM, Langley SA, Kennedy C, Costes SV, Karpen GH. Drosophila histone demethylase KDM4A has enzymatic and non-enzymatic roles in controlling heterochromatin integrity. Dev Cell. 2017; 42(2):156–169.e5.
crossref
36. Lorbeck MT, Singh N, Zervos A, Dhatta M, Lapchenko M, Yang C, et al. The histone demethylase Dmel\Kdm4A controls genes required for life span and male-specific sex determination in Drosophila. Gene. 2010; 450(1-2):8–17.
crossref
37. Fujiwara K, Fujita Y, Kasai A, Onaka Y, Hashimoto H, Okada H, et al. Deletion of JMJD2B in neurons leads to defective spine maturation, hyperactive behavior and memory deficits in mouse. Transl Psychiatry. 2016; 6(3):e766.
crossref
38. Zheng Y, Xue Y, Ren X, Liu M, Li X, Jia Y, et al. The lysine demethylase dKDM2 is non-essential for viability, but regulates circadian rhythms in Drosophila . Front Genet. 2018; 9:354.
crossref
TOOLS
ORCID iDs

Sung Yeon Park
https://orcid.org/0000-0002-2942-4101

Jieun Seo
https://orcid.org/0000-0003-1319-832X

Yang-Sook Chun
https://orcid.org/0000-0002-1261-9498

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