Dataset DOI: (https://doi.org/10.5061/dryad.brv15dvr6)

Description of the data and file structure

This dataset contains the original raw data supporting the manuscript entitled:

"Bmlebocin3 Could Inhibit Bombyx mori Cytoplasmic Polyhedrosis Virus in Silkworms via the IMD Pathway and Through the 20E Modulations."

The study investigates the antiviral function of Bombyx mori Lebocin-3 (BmLEB3) against Bombyx mori cytoplasmic polyhedrosis virus (BmCPV) and explores its regulation through the Immune Deficiency (IMD) signaling pathway and 20-hydroxyecdysone (20E).

The dataset contains the complete experimental data supporting the figures presented in the manuscript, including original RT-qPCR Ct values, molecular docking results, and uncropped Western blot images.

Dataset Organisation

The dataset is organised into three folders.

Folder 1 = RT-qPCR Raw Data

Description

This folder contains ten Microsoft Excel workbooks comprising the original RT-qPCR data generated during this study.

Workbooks 1–5:

These workbooks contain the original Ct values used to generate the main figures presented in the manuscript.

Workbook1 = Figure 1 complete

This workbook contains the original RT-qPCR Ct values used to generate Figure 1 of the manuscript.

Worksheet: Figure 1A

Original Ct values used to generate Figure 1A:

Treatment:

different body tissues (e.g., midgut, hemolymph, Malpighian tubules, etc.)

Control group:

fat body tissue

Worksheet: Figure 1B

Original Ct values used to generate Figure 1B.

Treatment group:

BmCPV-infected midgut tissue

Control group:

Uninfected midgut tissue (PBS-fed control)

Worksheet: Figure 1C

Original Ct values used to generate Figure 1C.

Treatment group:

BmCPV-infected BmN cells

Control group:

PBS-treated BmN cells

Workbook2 = Figure 2 complete

This workbook contains the original RT-qPCR Ct values used to generate Figure 2 of the manuscript.

Worksheet: Figure 2A

Ct values used to generate Figure 2A.

Treatment groups:

BmN cells treated with dsRNA targeting BmLEB3 at different concentrations:

1 µg dsRNA-BmLEB3

3 µg dsRNA-BmLEB3

5 µg dsRNA-BmLEB3

Control group:

BmN cells treated with dsRED

Worksheet2: Figure 2B

This worksheet contains the CT values used to generate Figure 2B

Treatment group:

BmN cells transfected with 5 µg dsRNA targeting BmLEB3

Control group:

BmN cells transfected with control dsRED

Experimental variable:

Different post-transfection time points

Worksheet: Figure 2C

This worksheet contains the CT values used to generate Figure 2C

Treatment group:

BmN cells transfected with dsRNA targeting BmLEB3 (dsBmLEB3) followed by BmCPV infection

Control group:

BmN cells transfected with dsRNA-dsRED followed by BmCPV infection

Experimental variable:

Different post-treatment time points after virus infection

Workbook3 = Figure 3 Complete

The dataset includes Ct values used to generate the complete Figure 3

worksheet: Figure 3B

This worksheet contains the CT values used to generate Figure 3B

Treatment group:

Cells transfected with pIEX-BmLEB3-V5 overexpression plasmid, followed by BmCPV infection

Control group:

Cells transfected with empty vector (pIEX), followed by BmCPV infection

Objective:

To evaluate the effect of BmLEB3 overexpression on viral infection and replication

Workbook: Figure 4 Complete

The dataset includes Ct values used to generate the complete Figure 4

Worksheet: Figure 4A

The worksheet contains CT values used to generate Figure 4A

Treatment groups:

BmN cells treated with different concentrations of 20-hydroxyecdysone (20E)

Control group:

0.025% DMSO-treated BmN cells

Time point:

Cells were collected at 24 hours post-treatment

Objective:

To evaluate the effect of 20E treatment on BmLEB3 in BmN cells.

Worksheet: Figure 4B

The worksheet contains the CT values used to generate Figure 4B

Treatment group:

BmN cells treated with 4 µM 20E

Control group:

BmN cells treated with 0.025% DMSO

Experimental variable:

Cells were collected at different time points after treatment

Objective:

To evaluate the time-dependent effect of 20E on BmLEB3 in BmN cells

Worksheet: 4C

The worksheet contains the CT values used to generate Figure 4C

Treatment group:

BmN cells treated with 4 µM 20E and infected with BmCPV

Control group:

BmN cells treated with 0.025% DMSO and infected with BmCPV

Experimental variable:

Cells were collected at different post-treatment time points

Objective:

To evaluate the effect of 20E on BmCPV replication through increased BmLEB3 expression.

Worksheet: Figure 4D

The worksheet contains the CT values used to generate Figure 4D

Treatment group:

Midgut tissues treated with 20-hydroxyecdysone (20E)

Dose:

4 µL of 2.4 µg/µL 20E solution

Control group:

DMSO injected midgut tissues

Experimental variable:

Samples collected at different time points after treatment

Objective:

To evaluate the effect of 20E on BmLEB3 expression over time in silkworm midgut tissue

Worksheet: Figure 4E

The worksheet contains the CT values used to generate Figure 4E

Treatment group:

Midgut tissues treated with 20E (4 µL of 2.4 µg/µL solution) and infected with BmCPV

Control group:

Midgut tissues treated with DMSO and infected with BmCPV

Experimental objective:

To evaluate the effect of 20E-induced upregulation of BmLEB3 on BmCPV replication

Workbook = Figure 5 Complete

The dataset includes Ct values used to generate the complete Figure 5

Worksheet: Figure 5A

The worksheet contains the CT values used to generate Figure 5A

Treatment group:

BmCPV-infected BmN cells

Control group:

Cells treated with PBS

Experimental variable:

Samples collected at different time points post-treatment/infection

Objective:

To evaluate the effect of BmCPV infection on BmPGRP-S1 expression

Worksheet: Figure 5B

The worksheet contains the CT values used to generate Figure 5B

Treatment group:

Bmn cells infected with BmCPV

Control group:

BmN cells treated with PBS

Experimental variable:

Samples collected at different time points post-infection

Objective:

To evaluate the effect of BmCPV infection on the expression of the BmRelish.

Worksheet: Figure 5C

The worksheet contains the CT values used to generate Figure 5C

Treatment group:

BmN cells treated with dsRNA targeting BmPGRP-S1

Control group:

BmN cells treated with dsRED

Experimental variable:

Samples collected at different time points after RNAi treatment

Objective:

To evaluate the effect of BmPGRP-S1 silencing on BmPGRPS1

Worksheet: Figure 5D

The worksheet contains the CT values used to generate Figure 5D

Treatment group:

BmN cells treated with dsRNA targeting BmPGRP-S1

Control group:

BmN cells treated with dsRED

Experimental variable:

Samples collected at different time points after RNAi treatment

Objective:

To evaluate the effect of BmPGRP-S1 silencing on BmRelish

Worksheet: Figure 5E

The worksheet contains the CT values used to generate Figure 5E

Treatment group:

BmN cells treated with dsRNA targeting BmPGRP-S1

Control group:

BmN cells treated with dsRED

Experimental variable:

Samples collected at different time points after RNAi treatment

Objective:

To evaluate the effect of BmPGRP-S1 silencing on BmLEB3

Worksheet: Figure 5F

The worksheet contains the CT values used to generate Figure 5F

Treatment group:

BmN cells treated with dsRNA targeting BmRelish

Control group:

BmN cells treated with dsRED

Experimental variable:

Samples collected at different time points after RNAi treatment

Objective:

To evaluate the effect of BmRelish silencing on BmRelish

Worksheet: Figure 5G

The worksheet contains the CT values used to generate Figure 5G

Treatment group:

BmN cells treated with dsRNA targeting BmRelish

Control group:

BmN cells treated with dsRED

Experimental variable:

Samples collected at different time points after RNAi treatment

Objective:

To evaluate the effect of BmRelish silencing on BmLEB3

Workbooks 6–10:

These workbooks contain the original CT and fluorescence intensity values used to generate the supplementary figures presented in the manuscript.

Workbook: Supplementary figure 3

This workbook contains the original values used to generate the supplementary Figure 3.

Worksheet = Supplementary Figure 3

This worksheet contains the original CCK-8 assay values used to analyze the effect of BmLEB3 knockdown on cell viability in BmN cells.

Treatment group:

BmN cells treated with 5 µg dsRNA targeting BmLEB3

Control group:

BmN cells treated with dsRED

Experimental variable:

Cells were collected at different time points after RNAi treatment

Objective:

To evaluate the effect of BmLEB3 silencing on cell viability and proliferation

Workbook: Supplementary Figure 4 Complete

The workbook contains CCk-8 values and CT values used to generate the supplementary Figure 4

Worksheet: Supplementary Figure 4B

This worksheet contains the original CCK-8 assay values used to analyse the effect of different concentrations of 20E on cell viability in BmN cells.

Treatment group:

BmN cells treated with different concentrations of 20E

Control group:

BmN cells treated with 0.025%

Experimental variable:

Cells were collected 24 hours after treatment with 20E

Objective:

To evaluate the effect of 20 treatments on cell viability and proliferation

Worksheet: Supplementary Figure 4C

The worksheet contains the CT values used to generate the supplementary Figure 4C

Treatment group:

BmN cells treated with different concentrations of 20E

Control group:

BmN cells treated with 0.025% DMSO

Experimental variable:

Cells were collected 24 hours after treatment with 20E

Objective:

To evaluate the effect of different concentrations of 20E on BmLEB3 expression in BmN cells

Workbook = Supplementary Figure 5 complete

The workbook contains CT values used to generate the supplementary Figure 5

Worksheet: Supplementary Figure 5A, B, and C

The worksheet contains the CT values used to generate Figures 5A, 5B, and 5C

Treatment group:

BmN cells treated with 4µM 20E

Control group:

BmN cells treated with 0.025% DMSO

Experimental variable:

Cells were collected 24 hours after treatment with 20E

Objective:

To evaluate the effect of 20 treatments on the expression of different genes (BmBRcZ4, BmECR, and BmETS).

Workbook = Supplementary Figure 6

The workbook contains CT values used to generate the supplementary Figure 6

Worksheet: Supplementary Figure 6A,B, and C

The sheet contains CT values used to generate Supplementary Figures 6A, 6B, and 6C.

Treatment group:

midgut of the silkworms injected with 20E (4 µL of 2.4 µg/µL solution).

Control group:

midgut of the silkworms injected with DMSO.

Experimental variable:

Objective:

To evaluate the effect of 20 treatments on the expression of different genes (BmBRcZ4, BmECR, and BmETS).

Workbook = Supplementary Figure7

The workbook contains CT values used to generate the supplementary Figure 7

Worksheet: Supplementary Figure 7A

The worksheet contains Ct values used to generate Supplementary Figure 7A.

Treatment group:

BmN cells treated with 4µM 20E

Control group:

BmN cells treated with 0.025% DMSO

Experimental variable:

Samples collected at different time points after20E treatment

Objective:

To evaluate the effect of 20 treatments on the expression of BmPGRPS1.

Worksheet: Supplementary Figure 7B

The worksheet contains Ct values used to generate Supplementary Figure 7B.

Treatment group:

BmN cells treated with 4µM 20E

Control group:

BmN cells treated with 0.025% DMSO

Experimental variable:

Samples collected at different time points after 20E treatment

Objective:

To evaluate the effect of 20 treatments on the expression of BmRelish.

Folder 2:Raw Western Blot Images

Description

The folder includes raw, unprocessed Western blot images (TIFF format) used to prepare figures for the manuscript. These images represent the original experimental data before any cropping or layout adjustments.

These worksheets follow the same format as those used for the main figures.

Panel descriptions

Panel A

Original uncropped immunoblot used to detect V5-tagged BmLEB3 in BmN cells transfected with pIEX-BmLEB3-V5. Protein expression was examined at different post-transfection time points using an anti-V5 antibody (1:2000 dilution).

Panel B

Original uncropped immunoblot of the empty pIEX vector control collected at the corresponding time points. This panel serves as the negative control for the BmLEB3 overexpression experiment.

Panel C

Original uncropped immunoblot of α-tubulin, used as the loading control for the corresponding samples

Purpose

These uncropped Western blot images are provided as the original experimental data.

File format

TIFF (.tif)

Folder 3: Molecular Docking Data

Description

The Folder contains raw molecular docking data analysing the interaction between 20-hydroxyecdysone (20E) and BmPGRP-S1 using MOE software, including docking output files and visualisation data.

Figure 6 docking:

File description

Docking Pose 1

Three-dimensional visualization of the docked BmPGRP-S1–20E complex viewed from the first orientation.

Docking Pose 2

Three-dimensional visualization of the same BmPGRP-S1–20E docking complex viewed from a different angle.

Docking Pose 3

Three-dimensional visualization of the same docking complex from an alternative perspective, highlighting the spatial arrangement of the ligand within the predicted binding pocket.

Docking Pose 4

Three-dimensional visualization of the same docking complex viewed from another angle, providing a comprehensive representation of the protein–ligand interaction.

BmPGRPS1+20E Interaction. moe

Original MOE project file containing the complete molecular docking analysis, including the docked complex, docking scores, binding poses, and interaction information between BmPGRP-S1 and 20-hydroxyecdysone (20E).

BmPGRPS1+20E Interaction Pose 1.mdb

MOE database file containing the docking coordinates and interaction data for the highest-ranked binding pose (Pose 1).

Ligand Interaction.jpeg

Two-dimensional interaction diagram showing the predicted interactions between BmPGRP-S1 and 20-hydroxyecdysone (20E), including hydrogen bonds, hydrophobic interactions, and interacting amino acid residues.

Ligand Interaction.txt

Microsoft Word document containing the annotated ligand–protein interaction analysis and the corresponding two-dimensional interaction diagram.

Software/Codes

Software required includes Microsoft Excel for RT-qPCR data, MOE for molecular docking files, and standard image viewers for TIFF Western blot images