Dataset DOI: 10.5061/dryad.8sf7m0d36

Description of the data and file structure

The data include pupal size of fed and starved male and female flies across 196 lineages, the SNP identities for those lineages at 4,438,427 loci, the annotations of those SNPs, the code to conduct an MLM GWAS on those SNPs and a subsequent VEGAS, the results of the GWAS and VEGAS, the data for the functional validation of candidate genes from the MLM GWAS and the code to analyze it.

Files and variables

File: Analysis__Enrichment.Rmd

Description: Code to perform enrichment analysis (GO, KEGG) on MLM GWAS hits, as well as permutation tests and VEGAS to assess aggregate association signals between growth-regulatory or nutrient-signaling genes and variation in SSP, SSD, and body size.

File: Analysis_MLM_GWAS.Rmd

Description: Code to perform a Mixed Linear Model Genome-Wide Association Study on body size traits.

File: Figure_1_and_S1.Rmd

Description: Code to generate the charts in Figures 1 and S1 of the manuscript.

File: Analysis_VEGAS.Rmd

Description: Code to conduct a Versatile Gene-based Association Study (VEGAS) on body size traits using the results from the MLM GWAS.

File: plink.eigenvec

Description: The first 20 principle components of the genetic distance matrix for the DGRP lineages.

File: focal_growth_genes.csv

Description: List of 330 focal growth genes

Variables

• FBgn: Flybase gene number
• CG: Computed gene identifier
• gene: Gene symbol
• name: Gene name
• EntrezID: Entrex identifier

File: Figure_2.Rmd

Description: Code to generate the charts in Figure 2 of the manuscript.

File: DGRP_pupa.csv

Description: Phenotype data

Variables

• id: Unique sample identifier
• line: DRPG lineage
• block: Experimental block
• day: Day of starvation. D0 = fed, D1 = starved
• sex: Male or female
• pupa: Log pupal area (µm²)

File: Wolb_and_Inv_Status.csv

Description: Lineage status for Wolbachia infection and inversions.

Variables

• DGRP Line: DGRP lineage 
• Infection Status: Wolbachia infection status (y/n)
• In.2L.t: Inversion status (ST = standard, INV = inversion. ST/INV = heterozygous for the inversion)
• In.2R.NS: As above
• In.2R.Y1: As above
• In.2R.Y2: As above
• In.2R.Y3: As above
• In.2R.Y4: As above
• In.2R.Y5: As above
• In.2R.Y6: As above
• In.2R.Y7: As above
• In.3L.P: As above
• In.3L.M: As above
• In.3L.Y: As above
• In.3R.P: As above
• In.3R.K: As above
• In.3R.Mo: As above
• In.3R.C: As above

File: IIS_genes.csv

Description: List of 70 focal nutrient-signaling genes

Variables

• FBgn: Flybase gene number
• CG: Computed gene identifier
• gene: Gene symbol
• name: Gene name
• EntrezID: Entrez identifier

File: dgrp2.tgeno

Description: SNP identities for DGRP lineages

Variables

• chr: Chromosome arm on which the variant is located (e.g., 2L, 2R, 3L, 3R, X).
• pos: Genomic position of the variant (base-pair coordinate on the given chromosome).
• id: Variant identifier, typically combining chromosome, position, and variant type (e.g., “2L_4998_SNP”).
• ref: Reference allele at this genomic position (as in the reference genome).
• alt: Alternate (non-reference) allele at this position.
• refc: Number of sequencing reads supporting the reference allele across all lines/sample(s) used for the variant call.
• altc: Number of sequencing reads supporting the alternate allele across all lines/sample(s) used for the variant call.
• qual: Overall quality score of the variant call (higher values indicate greater confidence).
• cov: Number of lines with non-missing genotype calls for this variant.
• line_21, line_26, line_28, …: Genotype call for the line at this variant, coded as:
  • 0: Homozygous for the reference allele
  • 2: Homozygous for the alternate allele
  • -: Missing or no reliable genotype call

File: Pvalues_annot_Column_Descriptions.txt

Description: Column description for Pvalues_annot data table

File: dgrp.fb549.annot.csv

Description: SNP annotations based on FlyBase release 5.49

Variables:

• ID: SNP identifier
• annot: SNP annotation

File: DGRP2.gwas.BSD0.all.assoc

Description: DGPR2 GWAS results for fed body size

Variables:

• ID: Unique identifier for each SNP, combining chromosome, genomic position, and variant type (e.g., “2L_5317_SNP”).
• MinorAllele: Allele with the lower frequency in the sampled population.
• MajorAllele: Allele with the higher frequency in the sampled population.
• MAF: Minor Allele Frequency — the proportion of chromosomes carrying the minor allele.
• MinorAlleleCount:Total number of observed copies of the minor allele in the dataset.
• MajorAlleleCount: Total number of observed copies of the major allele in the dataset.
• FemalePval: P-value from the single-locus association test (standard linear model) on females only.
• FemaleMixedPval: P-value from the mixed-model association test (MLM/linear mixed model), which accounts for relatedness or population structure, on females only.
• MalePval: P-value from the single-locus association test (standard linear model) on males only.
• MaleMixedPval: P-value from the mixed-model association test (MLM/linear mixed model), which accounts for relatedness or population structure, on males only.
• AvgPval:  P-value from the single-locus association test (standard linear model) on the average of male and females.
• AvgMixedPval: P-value from the mixed-model association test (MLM/linear mixed model), which accounts for relatedness or population structure, on average of male and females.
• DiffPval: P-value from the single-locus association test (standard linear model) on absolute differences between male and female (SSD).
• DiffMixedPval: P-value from the mixed-model association test (MLM/linear mixed model), which accounts for relatedness or population structure, on absolute differences between male and female (SSD).

File: DGRP2.gwas.SSD0.all.assoc

Description: DGPR2 GWAS results for fed SSD

Variables:

• ID: Unique identifier for each SNP, combining chromosome, position, and variant type (e.g., “2L_5317_SNP”).
• MinorAllele: The allele with the lower frequency in the sampled population.
• MajorAllele: The allele with the higher frequency in the sampled population.
• MAF: Minor Allele Frequency — the proportion of chromosomes carrying the minor allele.
• MinorAlleleCount: Total number of observed copies of the minor allele across all genotyped samples.
• MajorAlleleCount: Total number of observed copies of the major allele across all genotyped samples.
• SinglePval: P-value from the single-locus association test (standard linear model).
• SingleMixedPval: P-value from the mixed-model association test (MLM/linear mixed model), which accounts for relatedness or population structure.

File: DGRP2.gwas.BSD1.all.assoc

Description: DGPR2 GWAS results for starved  body size

Variables:

• ID: Unique identifier for each SNP, combining chromosome, genomic position, and variant type (e.g., “2L_5317_SNP”).
• MinorAllele: Allele with the lower frequency in the sampled population.
• MajorAllele: Allele with the higher frequency in the sampled population.
• MAF: Minor Allele Frequency — the proportion of chromosomes carrying the minor allele.
• MinorAlleleCount:Total number of observed copies of the minor allele in the dataset.
• MajorAlleleCount: Total number of observed copies of the major allele in the dataset.
• FemalePval: P-value from the single-locus association test (standard linear model) on females only
• FemaleMixedPval: P-value from the mixed-model association test (MLM/linear mixed model), which accounts for relatedness or population structure, on females only
• MalePval: P-value from the single-locus association test (standard linear model) on males only
• MaleMixedPval: P-value from the mixed-model association test (MLM/linear mixed model), which accounts for relatedness or population structure, on males only
• AvgPval:  P-value from the single-locus association test (standard linear model) on average of male and females
• AvgMixedPval: P-value from the mixed-model association test (MLM/linear mixed model), which accounts for relatedness or population structure, on average of male and females.
• DiffPval: P-value from the single-locus association test (standard linear model) on absolute differences between male and female (SSD).
• DiffMixedPval: P-value from the mixed-model association test (MLM/linear mixed model), which accounts for relatedness or population structure, on absolute differences between male and female (SSD).

File: DGRP2.gwas.SSD1.all.assoc

Description: DGPR2 GWAS results for starved SSD

Variables:

• ID: Unique identifier for each SNP, combining chromosome, position, and variant type (e.g., “2L_5317_SNP”).
• MinorAllele: The allele with the lower frequency in the sampled population.
• MajorAllele: The allele with the higher frequency in the sampled population.
• MAF: Minor Allele Frequency — the proportion of chromosomes carrying the minor allele.
• MinorAlleleCount: Total number of observed copies of the minor allele across all genotyped samples.
• MajorAlleleCount: Total number of observed copies of the major allele across all genotyped samples.
• SinglePval: P-value from the single-locus association test (standard linear model).
• SingleMixedPval: P-value from the mixed-model association test (MLM/linear mixed model), which accounts for relatedness or population structure.

File: DGRP2.gwas.SSP_indep.all.assoc

Description: DGPR2 GWAS results for nutritional plasticity of body size, independent of fed body size.

Variables:

• ID: Unique identifier for each SNP, combining chromosome, genomic position, and variant type (e.g., “2L_5317_SNP”).
• MinorAllele: Allele with the lower frequency in the sampled population.
• MajorAllele: Allele with the higher frequency in the sampled population.
• MAF: Minor Allele Frequency — the proportion of chromosomes carrying the minor allele.
• MinorAlleleCount:Total number of observed copies of the minor allele in the dataset.
• MajorAlleleCount: Total number of observed copies of the major allele in the dataset.
• FemalePval: P-value from the single-locus association test (standard linear model) on females only.
• FemaleMixedPval: P-value from the mixed-model association test (MLM/linear mixed model), which accounts for relatedness or population structure, on females only.
• MalePval: P-value from the single-locus association test (standard linear model) on males only.
• MaleMixedPval: P-value from the mixed-model association test (MLM/linear mixed model), which accounts for relatedness or population structure, on males only.
• AvgPval:  P-value from the single-locus association test (standard linear model) on average of male and females.
• AvgMixedPval: P-value from the mixed-model association test (MLM/linear mixed model), which accounts for relatedness or population structure, on average of male and females.
• DiffPval: P-value from the single-locus association test (standard linear model) on absolute differences between male and female (SSP).
• DiffMixedPval: P-value from the mixed-model association test (MLM/linear mixed model), which accounts for relatedness or population structure, on absolute differences between male and female (SSP).

File: DGRP2.gwas.SSP.all.assoc

Description: DGPR2 GWAS results for nutritional plasticity of body size.

Variables:

• ID: Unique identifier for each SNP, combining chromosome, genomic position, and variant type (e.g., “2L_5317_SNP”).
• MinorAllele: Allele with the lower frequency in the sampled population.
• MajorAllele: Allele with the higher frequency in the sampled population.
• MAF: Minor Allele Frequency — the proportion of chromosomes carrying the minor allele.
• MinorAlleleCount:Total number of observed copies of the minor allele in the dataset.
• MajorAlleleCount: Total number of observed copies of the major allele in the dataset.
• FemalePval: P-value from the single-locus association test (standard linear model) on females only.
• FemaleMixedPval: P-value from the mixed-model association test (MLM/linear mixed model), which accounts for relatedness or population structure, on females only.
• MalePval: P-value from the single-locus association test (standard linear model) on males only.
• MaleMixedPval: P-value from the mixed-model association test (MLM/linear mixed model), which accounts for relatedness or population structure, on males only.
• AvgPval:  P-value from the single-locus association test (standard linear model) on average of male and females.
• AvgMixedPval: P-value from the mixed-model association test (MLM/linear mixed model), which accounts for relatedness or population structure, on average of male and females.
• DiffPval: P-value from the single-locus association test (standard linear model) on absolute differences between male and female (SSP).
• DiffMixedPval: P-value from the mixed-model association test (MLM/linear mixed model), which accounts for relatedness or population structure, on absolute differences between male and female (SSP).

File: Pvalues_annot.csv

Description: Complete DGRP2 GWAS, MLM GWAS and VEGAS results

Variables

• ID: SNP ID
• fbgn: Flybase gene number
• CG: Computed gene identifier
• entrez: Entrez identifier
• gene: Gene symbol
• fullname: Full gene name
• L_FBSD0: p-value for MLM GWAS on fed female body size
• L_MBSD0: p-value for MLM GWAS on fed male body size
• L_AvgBSD0: p-value for MLM GWAS on fed sex-averaged body size
• L_SSD0: p-value for MLM GWAS on fed SSD
• L_FBSD1: p-value for MLM GWAS on starved female body size
• L_MBSD1: p-value for MLM GWAS on starved male body size
• L_AvgBSD1: p-value for MLM GWAS on starved sex-averaged body size
• L_SSD1: p-value for MLM GWAS on starved SSD
• L_FPlast: p-value for MLM GWAS on female plasticity
• L_MPlast: p-value for MLM GWAS on male plasticity
• L_AvgPlast: p-value for MLM GWAS on sex-averaged plasticity
• L_SSP_ALL: p-value for MLM GWAS on SSP (both SSP and SSP independent of body size)
• V_FBSD0: p-value for VEGAS on fed female body size
• V_MBSD0: p-value for VEGAS on fed male body size
• V_AvgBSD0: p-value for VEGAS on fed sex-averaged body size
• V_SSD0:  p-value for VEGAS on fed SSD
• V_FBSD1: p-value for VEGAS on starved female body size
• V_MBSD1: p-value for VEGAS on starved male body size
• V_AvgBSD1: p-value for VEGAS on starved sex-averaged body size
• V_SSD1: p-value for VEGAS on starved SSD
• V_FPlast: p-value for VEGAS on female plasticity
• V_MPlast: p-value for VEGAS on male plasticity
• V_AvgPlast: p-value for VEGAS on sex-averaged plasticity
• V_SSP_ALL: p-value for VEGAS on SSP (both SSP and SSP independent of body size)
• FBSD0: p-value for DGRP2 GWAS on fed female body size
• MBSD0: p-value for DGRP2 GWAS on fed male body size
• AvgBSD0: p-value for DGRP2 GWAS on fed sex-averaged body size
• SSD0: p-value for DGRP2 GWAS on fed SSD
• FBSD1: p-value for DGRP2 GWAS on starved female body size
• MBSD1: p-value for DGRP2 GWAS on starved male body size
• AvgBSD1: p-value for DGRP2 GWAS on starved sex-averaged body size
• SSD1: p-value for DGRP2 GWAS on starved SSD
• FPlast: p-value for DGRP2 GWAS on female plasticity
• MPlast: p-value for DGRP2 GWAS on male plasticity
• AvgPlast: p-value for DGRP2 GWAS on sex-averaged plasticity
• SSP: p-value for DGRP2 GWAS on SSP
• SSP_indep: p-value for DGRP2 GWAS on SSP independent of fed body size
• annot: Full SNP annotation from FlyBase release 5.49

File: Validation.csv

Description: Results of functional analysis on candidate genes that affect SSP

Variables

• gene: Target gene
• pupa: Log pupal area (µm²)
• sex: Sex
• diet: Fed 
• type: Day of starvation. D0 = fed, D1 = starved

Code/software

All the code can be run in R version 4.5.1 (2025-06-13).