Journal: Cellular and molecular life sciences : CMLS

Article Title: Dnajc5b contributes to male fertility by maintaining the mitochondrial functions and autophagic homeostasis during spermiogenesis.

doi: 10.1007/s00018-024-05552-1

Figure Lengend Snippet: Fig. 1 Meta-analysis shows Dnajc5b is a spermatogenesis- associated gene. a Workflow of data collection, meta-analysis, and candidate genes mining. b The age distribution of porcine samples. n = 11 (IMM, 7 samples have exact age information, and 4 samples are in infancy) and 59 (MAT). pnd: postnatal days. c The age distribution of murine samples. n = 67 (IMM) and 48 (MAT). pnd: postnatal days. d The overlapping DEGs in pig and mouse. There are 438 overlap ping DEGs enriched in MAT groups and 50 overlapping DEGs enriched in IMM groups. e, f The biological process (e) and cellular component (f) enrichment analy sis of 438 MAT-enriched genes from plot d. g The fold change values and effect sizes distribu tion of all pig genes. h The fold change values and effect sizes distribution of all mouse genes. i The top 10 MAT-enriched genes ranked by combined effect sizes of 488 murine DEGs. The “PFS” (0–1) was calculated by Sper matogenesisOnline1.0, reflecting the probability of gene function in spermatogenesis

Article Snippet: The primary antibodies included DNAJC5B (1:1000, Proteintech, 17364-1-AP), P62 (1:1000, Zen-Bio, 380612), LC3B (1:2000, Abclonal, A7198), FBXL4 (1:1000, Zen-Bio, 863894), LONP1 (1:1000, Abclonal, A4293), ATP1A1 (1:5000, Abmart, T55159), ATF5 (1:1000, Abclonal, A3563), ACTB (1:10000, Abclonal, AC026), GRP75 (1:1000, Abclonal, A11256), α- TUBULIN (1:5000, Abclonal, AC012), GAPDH (1:10000, Abclonal, AC033), ODF1 (1:2000, Abcam, ab197029), and FLAG (1:3500, Abmart, M20008).

Techniques:

Journal: Cellular and molecular life sciences : CMLS

Article Title: Dnajc5b contributes to male fertility by maintaining the mitochondrial functions and autophagic homeostasis during spermiogenesis.

doi: 10.1007/s00018-024-05552-1

Figure Lengend Snippet: Fig. 5 Loss of Dnajc5b breaks mitochondrial function and activates autophagy in the testis. a TEM observa tion of the control and CKO testicular spermatids. The red arrows indicate the condensed mitochondria and the red arrowheads indicate the mitochondria with loose matrix. The cartoon models show the condensed mito chondria in control spermatids and the mitochondria with loose matrix in CKO spermatids; the mitochon drial matrix is shown in yellow and the mitochondrial intermembrane space is shown in green. nu: nucleus; acr: acrosome. Scale bars, 1 μm. b The ROS levels of the control and CKO testicular cells. n = 5. c, d The mRNA levels of the genes related to unfolding protein response of mitochondria (UPRmt, c) and endoplas mic reticulum (UPRER, d) in control and CKO testes by qRT-PCR. n = 5. e Western blot of UPRmt proteins (ATF5 and LONP1) and autophagy-related proteins (LC3B and P62) in control and CKO testes. f Quantita tive results of protein expression levels in plot e. n = 3. g Immunofluorescence of LC3B in control and CKO testes. The autophagosomes are indicated by LC3B puncta. Scale bars, 50 μm. h The ratio of LC3B puncta per testicular cell. Random areas were selected for counting the number of LC3B puncta. SPg: spermato gonia; SPc: spermatocyte; RS: round spermatid; ES: elongating spermatid; n = 10. i Western blot of LC3B-II and LC3B-I in testicular mitochondria and cytoplasm. CPT1A is the marker of mitochondria. α-TUBULIN is the marker of cytoplasm. Data are represented as mean ± SEM of three independent experiments

Article Snippet: The primary antibodies included DNAJC5B (1:1000, Proteintech, 17364-1-AP), P62 (1:1000, Zen-Bio, 380612), LC3B (1:2000, Abclonal, A7198), FBXL4 (1:1000, Zen-Bio, 863894), LONP1 (1:1000, Abclonal, A4293), ATP1A1 (1:5000, Abmart, T55159), ATF5 (1:1000, Abclonal, A3563), ACTB (1:10000, Abclonal, AC026), GRP75 (1:1000, Abclonal, A11256), α- TUBULIN (1:5000, Abclonal, AC012), GAPDH (1:10000, Abclonal, AC033), ODF1 (1:2000, Abcam, ab197029), and FLAG (1:3500, Abmart, M20008).

Techniques: Control, Quantitative RT-PCR, Western Blot, Expressing, Immunofluorescence, Marker