Tau reduction impairs nephrocyte function in Drosophila

Abstract

Tau, a microtubule-associated protein, is known for its significant involvement in neurodegenerative diseases. While various molecular and immunohistochemical techniques have confirmed the presence of Tau in podocytes, its precise function within these cells remains elusive. In this study, we investigate the role of Tau in kidney podocytes using Drosophila pericardial nephrocytes as a model. We found that knockdown of Drosophila Tau in nephrocytes resulted in apoptotic cell death and the disruption of nephrocyte structure. Furthermore, we observed that decreased Tau levels induced genomic damage and abnormal distribution of γ-H2Av, altering nuclei architecture in nephrocytes, and affecting the nuclear membrane structure by interfering with lamin with aging. Additionally, Tau knockdown led to a reduction in lipid droplets in Drosophila fat body tissues, suggesting a potential role of Tau in inter-organ communication. These findings underscore the importance of Tau in the nephrocytes of Drosophila, and advocate further research to broaden our understanding of podocyte biology in kidney diseases. [BMB Reports 2025; 58(4): 169-174].

Fig. 1

Knockdown of dTau disrupts nephrocyte function in Drosophila. (A) Pericardial nephrocytes in the abdomen from mCD8−RFP-expressing flies under the control of snsGCN−Gal4. The RNAi-mediated knockdown of dTau caused severe morphological abnormalities in Drosophila nephrocytes (arrowheads). (B) The RNAi-mediated genetic knockdown of dTau in nephrocytes decreased lifespan (n > 150 each). (C) Pericardial nephrocytes (red) in the abdomen from mCD8−RFP-expressing flies under the control of snsGCN−Gal4. Nephrocytes dissected from (3 or 20)-d-old dTau RNAi-expressing flies were stained with anti-cleaved DCP−1 antibody (green) and DAPI (blue). Increased DCP−1 signals, a marker for apoptosis, were observed in the nephrocytes of dTau RNAi-expressing flies. (D) Quantification of the number of DCP−1 positive puncta in the nephrocytes of adult flies at (3 and 20)-d-old. A significant increase in the number of puncta was observed in the dTau RNAi-expressing nephrocytes, compared to the control. Error bars represent the mean ± standard deviation (n ≥ 10 nephrocytes for each genotype). Statistical significance was determined using Student’s t-test (*P < 0.05, ***P < 0.001). (E) Photomicrograph showing ingested AgNO₃ sequestered in larval nephrocytes. The pericardial nephrocytes in control flies contained silver, but not when dTau had been silenced (brown pigment). Dashed lines indicate the borders of pericardial nephrocytes. (F) AgNO₃ levels were determined from photomicrographs, and expressed relative to control nephrocytes in snsGCN−Gal4 transgenic flies. For quantification, 20 nephrocytes were analyzed from each of three female flies per genotype. Experimental significance was determined using Student’s t‐test (***P < 0.001).

Fig. 2

Knockdown of dTau disrupts nuclear structure and forms senescence-associated heterochromatin foci (SAHF) in nephrocytes. (A) Nephrocytes were stained with DAPI (blue) to visualize DNA. SAHF, a marker of cellular senescence, was observed in the nephrocytes of dTau RNAi-expressing flies (arrowheads). (B) Quantification of DAPI foci-positive nephrocytes in adult flies. A significant increase in the number of DAPI foci-positive nephrocytes was observed in the dTau RNAi-expressing nephrocytes, compared to the control. Error bars represent the mean ± standard deviation (n = 4 adult flies for each genotype). Statistical significance was determined using Student’s t-test (***P < 0.001). (C) Pericardial nephrocytes (red) in the abdomen of mCD8−RFP-expressing flies under the control of snsGCN−Gal4. Nephrocytes dissected from (3 or 20)-d-old dTau RNAi-expressing flies were stained with anti-histone H2Av antibody (green) and DAPI (blue). dTau knockdown exhibits increased frequencies of γ–H2Av foci in the nephrocytes of 3-d-old flies (arrowheads). Dispersed γ–H2Av foci in 20-d-old dTau RNAi-expressing flies were detected in the cytoplasm of nephrocytes. (D, E) Quantification of nuclear and cytoplasmic γ–H2Av fluorescent signal in the nephrocytes. Error bars represent the mean ± standard deviation (n ≥ 8 nephrocytes for each genotype). Statistical significance was determined using Student’s t-test (**P < 0.01, ***P < 0.001).

Fig. 3

Knockdown of dTau causes disrupted nuclear morphology by regulating lamin in nephrocytes. (A) The pericardial nephrocytes (red) in the abdomen from mCD8−RFP-expressing flies under the control of snsGCN−Gal4. Nephrocytes dissected from (3 or 20)-d-old dTau RNAi-expressing flies were stained with anti-lamin B antibody (green) and DAPI (blue). Abnormal nuclear morphology was observed in the nephrocytes of 3-d-old dTau RNAi-expressing flies (arrowheads). Knockdown of dTau caused loss of lamin in the nephrocytes of 20-d-old flies (yellow arrowheads). (B) Quantification of the nuclear circularity in the nephrocytes of adult flies at 3-d-old. Error bars represent the mean ± standard deviation (n ≥ 9 nephrocytes for each genotype). Statistical significance was determined using Student’s t-test (***P < 0.001).

Fig. 4

Nephrocyte-specific knockdown of dTau leads to smaller lipid droplets (LDs) in fat body tissues. (A) Body weight of dTau RNAi-expressing flies compared to control snsGCN−Gal4 flies. Error bars represent the mean ± standard deviation (n = 6 replicates; each replicate contained (7-10) flies). Experimental significance was determined using Student’s t‐test (***P < 0.001). (B) LDs labeled by Oil red O (red) in adult fat bodies from dTau RNAi-expressing flies. Knockdown of dTau exhibited lipid storage defects with small LDs (dotted box). (C) Graphs showing quantification of fluorescence intensity of LDs stained with Oil red O. Error bars represent the mean ± standard deviation (n = 7 abdomens for each genotype). Experimental significance was determined using Student’s t‐test (***P < 0.001). (D) LDs labeled by Oil red O (red) in larval fat bodies from dTau RNAi-expressing flies. Knockdown of dTau exhibited a small size of LDs. Dashed lines indicate the borders of cells. (E) Quantification of LD diameter in (D). Quantification of the diameter of the three largest LDs per cell in 10 cells from each genotype. Error bars represent the mean ± standard deviation. Experimental significance was determined using Student’s t‐test (***P < 0.001).