Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: Splicing factor Sf3b1 facilitates maintenance of neuronal dendrites by modulating mitochondrial health

doi: 10.1007/s00018-025-05860-0

Figure Lengend Snippet: Reduced growth of dendritic arbors upon depletion of Sf3b1 in ddaC and ddaF neurons. ( A ) Schematics showing three DrosDel lines, Df(2L)ED94 , Df(2L)ED49, and Df(2L)ED62 , used for lethality mapping in which deleted regions are filled with red dots, the genomic locations of Sf3b1 and lectin-24Db in the chromosomal 2L region are labeled by black boxes, transcripts are represented by blue bars, exons are represented by yellow bars, and untranslated regions are shown by gray bars, with arrows indicating the direction of transcription. The lectin-24Db MI11191 line carries a MiMIC insertion (black diamond) at the lectin-24Db locus and a deleted region (black dotted) encompassing Sf3b1 . The newly isolated Sf3b1 mt allele is also shown. ( B-E ) Images display MARCM clones of ddaC neurons for the ( B ) control ( GAL4 5-40 UAS-Venus::pm SOP-FLP #42 ; Tub-GAL80 FRT40A/FRT40A ), ( C ) Sf3b1 mt mutant, ( D ) Sf3b1 mt mutant carrying UAS-Sf3b1, and ( E ) Sf3b1 mt mutant carrying UAS-SF3B1 Lines. Scale bars are 50 μm. ( F ) Quantification of dendritic arbor terminals. ( G-I ) Images show MARCM clones of ddaF neurons for the ( G ) control, ( H ) Sf3b1 mt mutant, and ( I ) Sf3b1 mt mutant carrying UAS-Sf3b1 lines . Boxed regions are magnified underneath to show spike protrusions. Scale bars are 50 μm. ( J ) Quantification of dendritic arbor terminals. In ( F, J ), bar graphs represent the mean ± SEM, and comparisons between data groups are indicated by lines with statistical significance tested by Student’s t -test and shown as **** for p < 0.0001

Article Snippet: SF3B1 plasmids , Addgene [ ] , # 82576.

Techniques: Labeling, Isolation, Clone Assay, Control, Mutagenesis

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: Splicing factor Sf3b1 facilitates maintenance of neuronal dendrites by modulating mitochondrial health

doi: 10.1007/s00018-025-05860-0

Figure Lengend Snippet: Retracted branch growth in Sf3b1 knockdown neurons at the late larval stage. ( A ) Images display ddaC neurons from ppk-GAL4 -driven lacZ control and Sf3b1 RNAi transgenic larvae at 72, 90 and 120 h AEL, with dendrites marked by CD4-tdTom. Scale bars are 100 μm. ( B ) Quantification of dendritic terminal numbers. ( C ) Images of ddaC dendrites were acquired at 105 h AEL and 3 h later for the lacZ control and Sf3b1 RNAi knockdown lines. Changes in the length of terminal branches have been color-coded: newly emerged (red); vanished (blue); extended (pink); and retracted (cyan), all within the 3-h period. ( D ) Percentages of terminal branches exhibiting different dynamics (emerge, vanish, extend, and retract). In ( B, D ), bar ( B ) and dot ( D ) graphs show the mean ± SEM, and comparisons between two data groups were processed by Student’s t -test with ** for p < 0.01, *** for p < 0.001, **** for p < 0.0001, and ns for no significance ( p > 0.05)

Article Snippet: SF3B1 plasmids , Addgene [ ] , # 82576.

Techniques: Knockdown, Control, Transgenic Assay

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: Splicing factor Sf3b1 facilitates maintenance of neuronal dendrites by modulating mitochondrial health

doi: 10.1007/s00018-025-05860-0

Figure Lengend Snippet: Altered mRNA splicing events in Sf3b RNAi knockdown larvae. ( A ) Schematic showing genetic crosses of da-GAL4 females with w 1118 (control) or Sf3b RNAi males to produce larval offspring 24 h AEL for RNA sequencing and analyses. ( B ) The pie chart shows the result of rMATS analysis, with a threshold of |Δψ|> 0.1 and FDR ≤ 0.05, that identified 2,228 altered splicing events across 1,118 genes for the alternative 3’ splicing site (A3SS), alternative 5’ splicing site (A5SS), retained intron (RI), skipped-exon (SE), and mutually exclusive exon (MXE) categories in Sf3b1 RNAi knockdown larvae. Numbers of events (n) are displayed. The GO enrichment analysis of alternatively spliced genes identified from rMATS is shown at right. ( C ) Venn diagram depicting the 185 genes displaying intron splicing variants, 45 of which (3.6%) overlap with the 1,118 genes identified by rMATS. The GO enrichment analysis of genes exhibiting intron splicing variation, as derived by aligning CIGAR strings to the reference genome, is presented at right. The GO terms have been grouped into Molecular Function (MF), Cellular Component (CC), and Biological Process (BP). Circle size represents gene count, with darker colors indicating lower adjusted p-values (adj.p), as calculated using the Benjamini–Hochberg method, and denoting greater significance

Article Snippet: SF3B1 plasmids , Addgene [ ] , # 82576.

Techniques: Knockdown, Control, RNA Sequencing, Derivative Assay

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: Splicing factor Sf3b1 facilitates maintenance of neuronal dendrites by modulating mitochondrial health

doi: 10.1007/s00018-025-05860-0

Figure Lengend Snippet: Validation of altered mRNA splicing following Sf3b1 knockdown. ( A-D ) Integrative Genomics Viewer (IGV) was used to display base coverage in da-GAL4 control (red) and Sf3b1 RNAi knockdown (blue) conditions, with counts within circles indicating splice junction reads and the y-axis showing maximum exon coverage. ( A ) SE events for Sf3b3 and Opa1 . ( B ) RI events for TFAM and prel . ( C ) A5SS events for lap and Gptx . ( D ) A3SS events for hlk and msn . Gene schematics show the direction of transcription (black arrows), exons as blocks, introns as lines, and alternatively used coding regions in pale green. Exon numbers have been named according to the RA transcripts of Opa1 , TFAM , Gptx , hlk , and msn , and the RC transcript of the remaining genes. Gel images show amplified cDNA for control and Sf3b1 RNAi by RT-PCR with primers marked by red arrows and schematics for predicted splicing events at right. In addition to alternative splicing, other splicing variants in prel and msn were detected, including an unannotated retained intron in prel and an extra PCR product for msn (*)

Article Snippet: SF3B1 plasmids , Addgene [ ] , # 82576.

Techniques: Biomarker Discovery, Knockdown, Control, Amplification, Reverse Transcription Polymerase Chain Reaction, Alternative Splicing

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: Splicing factor Sf3b1 facilitates maintenance of neuronal dendrites by modulating mitochondrial health

doi: 10.1007/s00018-025-05860-0

Figure Lengend Snippet: Genes downregulated upon Sf3b1 RNAi knockdown are associated with mitochondrial functions. ( A, B ) GO enrichment analysis for significantly upregulated ( A ) or downregulated ( B ) DEGs following Sf3b1 RNAi knockdown. The GO terms have been categorized as Molecular Function (MF), Cellular Component (CC), or Biological Process (BP). Circle size reflects gene counts, with darker colors indicating lower adjusted p-values (adj.p) and denoting greater significance. ( C ) Volcano plot displaying significantly upregulated (log 2 FC > 1, red) and downregulated (log 2 FC < −1, blue) genes, both with [FDR] < 0.05. Those associated with mitochondrial functions, identified using the MitoMax database, are marked with darker dots, revealing a significant presence in the downregulated category (dark blue). ( D ) PANGEA network of enriched functional terms. The nodes represent enriched terms, emphasizing energy-associated processes among the downregulated mitochondrial-related genes highlighted in dark blue in ( C ). These include a."Oxidation–reduction process"(GO:0055114), b."Mitochondrial respiratory chain complex assembly"(GO:0033108), c."Electron transport chain"(GO:0022900), d."Mitochondrial translation"(GO:0032543), e."Fatty acid oxidation"(GO:0019395), f."Citrate cycle (TCA cycle)"(KEGG: map00020), g."Oxidative phosphorylation"(KEGG: map00190), and h."Complex I biogenesis"(Reactome: R-DME-6799198)

Article Snippet: SF3B1 plasmids , Addgene [ ] , # 82576.

Techniques: Knockdown, Functional Assay, Phospho-proteomics

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: Splicing factor Sf3b1 facilitates maintenance of neuronal dendrites by modulating mitochondrial health

doi: 10.1007/s00018-025-05860-0

Figure Lengend Snippet: Mitochondrial defects in Sf3b1 knockdown C4da neurons. ( A ) Images show Dendra2-labeled mitochondria in the cell bodies of C4da neurons from lacZ control and Sf3b1 RNAi knockdown lines driven by ppk-GAL4 , with enlarged views of yellow boxed regions at right. Scale bars are 5 μm. ( B ) Averaged mitochondrial length and area of neuronal cell bodies. ( C ) Images show the distribution of mitochondria in C4da neurons of lacZ control and Sf3b1 RNAi knockdown larvae at 120 h AEL, with Dendra2 labeling mitochondria and CD4-td-Tom labeling dendrites. Scale bars are 100 μm. Magnified views of selected boxed areas, showing combined images and individual images of dendrites (red) and mitochondria (green). ( D ) Quantification of mitochondrial density within major dendritic processes at 72, 90, and 120 h AEL. ( E ) Representative pseudo-color images (rainbow LUT, scale 0–1.6) illustrate ATP sensor intensity in lacZ control and Sf3b1 RNAi neurons following 1 mM Antimycin A treatment for 0, 5, 10, 20, and 30 min. Scale bar represents 10μm. ( F ) Line graph shows the iATPSnFR 1.0 :mRuby3 ratios in lacZ control (blue) and Sf3b1 RNAi (yellow) neurons following 1 mM Antimycin A treatment for 0, 5, 10, 20, and 30 min. The open circle represents C4da neuron ATP level in larvae dissected without Antimycin A treatment, incubated in HL3 medium for 60 min. Data represent mean ± SEM. Statistical significance between control and experimental groups at each indicated time point was determined using Student's t-test , * for p < 0.05 and ns, not significant. ( G ) Images show dendrites outlined by mRuby3 signals of the mRuby3-iATPSnFR 1.0 sensor driven by ppk-GAL4 in lacZ control and Sf3b1 RNAi knockdown larvae at 120 h AEL. Panels at right display the straightened dendritic processes (yellow lines in images at left) with images of mRuby3 (red), iATPSnFR 1.0 (green), and the iATPSnFR 1.0 :mRuby3 ratio pseudo-colored using a rainbow lookup table from 0–1. ( H ) Quantification of the iATPSnFR 1.0 :mRuby3 ratio. The bar graph shows the ratios for control and Sf3b1 RNAi dendrites at 72, 90, and 120 h AEL. Each data point represents the ratio from an individual segment mask. ( B, D, H ) Bar graphs show means ± SEM, with *** for p < 0.001, **** for p < 0.0001, and ns for no significance

Article Snippet: SF3B1 plasmids , Addgene [ ] , # 82576.

Techniques: Knockdown, Labeling, Control, Incubation

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: Splicing factor Sf3b1 facilitates maintenance of neuronal dendrites by modulating mitochondrial health

doi: 10.1007/s00018-025-05860-0

Figure Lengend Snippet: Rescue of impaired dendritic branching and diminished mitochondrial densities in Sf3b1 RNAi knockdown neurons. ( A ) Images display ppk-GAL4 -driven CD4-tdTom-labeled C4da neurons with Sf3b1 RNAi knockdown and co-expression of lacZ , AMPKα CA , Buffy , Lim3 , Pink1 , park , Nmnat , Marf , or PPP1R15 , Scale bars are 100 μm. ( B ) Quantification of terminal points in Sf3b1 RNAi knockdown neurons co-expressing one of the genes in (A) . ( C ) Mitochondrial distribution in Sf3b1 RNAi knockdown C4da neurons, as visualized using Dendra2 labeling for mitochondria and CD4-td-Tom for dendrites. Images show the effects of overexpressing lacZ , AMPKα CA , Pink1 , park , or Nmnat , driven by ppk-GAL4 . Scale bars are 100 μm. Enlarged boxed areas on right show merged and separate dendrite (red) and mitochondria (green) images. ( D ) Quantification of mitochondrial density in dendritic processes. In ( B, D ), bar graphs represent mean ± SEM. Statistical significance was determined relative to the lacZ control by Student’s t -test and is shown as *** for p < 0.001, **** for p < 0.0001, and ns for no significance

Article Snippet: SF3B1 plasmids , Addgene [ ] , # 82576.

Techniques: Knockdown, Labeling, Expressing, Control

Journal: Cellular and Molecular Life Sciences: CMLS

Article Title: Splicing factor Sf3b1 facilitates maintenance of neuronal dendrites by modulating mitochondrial health

doi: 10.1007/s00018-025-05860-0

Figure Lengend Snippet: Mitochondrial turnover in Sf3b1 RNAi neurons is partially restored by AMPKα CA , Pink1 , or park overexpression. ( A ) Images show merged green and red channels of MitoTimer-labeled mitochondria in the soma and dendritic processes (arrowheads mark the mitochondria in dendrites) in C4da neurons of lacZ control larvae, as well as for Sf3b1 RNAi larvae overexpressing lacZ , AMPKα CA , Pink1, or park . The bottom panel shows a pseudo-color representation of the red:green ratio ranging from 0–1. Scale bars are 5 μm. ( B ) Quantification of MitoTimer fluorescence ratios in the cell bodies (grey bars) and dendrites (pink bars) of C4da neurons. Each dot in the bar graph represents the red:green ratio for an individual mitochondrial ROI. Red/green ratios are expressed as percentages (raw ratio × 100). ( C ) Quantification of numbers of red puncta in the cell body. In ( B, C ), means ± SEM are shown. Statistical significance was determined by Student’s t -test, with comparisons between the indicated groups. Significance levels are denoted as * for p < 0.05, ** for p < 0.01, **** for p < 0.0001, and ns for no significance

Article Snippet: SF3B1 plasmids , Addgene [ ] , # 82576.

Techniques: Over Expression, Labeling, Control, Fluorescence