DOI: 10.5061/dryad.8sf7m0d36
title: Private QTLs underlie the genetic architecture of hierarchical size traits in <em>Drosophila</em>

description: Sex-specific plasticity (SSP) is the phenomenon whereby the size of one
 sex is more environmentally sensitive than the other, and is thought to
 underlie the developmental regulation and evolution of sexual size
 dimorphism (SSD). Sex-specific plasticity is a higher order phenotype that
 emerges due to the effect of the environment and sex on core growth
 regulatory mechanisms. Genetic variation in SSP necessarily requires sex-
 and environment-specific variation in growth, yet the
 developmental-genetic mechanisms enabling such context-dependent variation
 remain poorly understood. Using a genome-wide association study (GWAS) and
 functional validation in Drosophila melanogaster, we dissected the genetic
 architecture of body size, plasticity, SSD, and SSP across 196 isogenic
 lineages. We find that each phenotype is governed by largely
 non-overlapping sets of loci, with most candidate variants lying outside
 canonical growth pathways. Instead, size trait are shaped by “private
 QTLs” whose effects are limited to specific sex, trait, or environmental
 contexts. Functional knockdown of selected candidate genes for SSP
 revealed that while most did not affect SSP directly, many influenced body
 size, SSD, and plasticity, consistent with their nested phenotypic
 relationships. Together, our results suggest that context-dependent
 alleles in genes peripheral to core growth regulatory pathways drive
 variation in SSD and SSP, offering a mechanistic explanation for their
 evolutionary lability and highlighting the role of private QTLs in
 structuring complex trait architecture. The deposited data include the
 size phenotypes (196 lineages, female and male body size in fed and
 starved flies), the SNP identities per lineage, the eigen vectors for the
 population strutcture, the Wolbachia and inversion status of the lineages,
 the phenotype data for the functional analysis of candidate QTLs, the list
 of core growth-regulatory and nutrional-signal genes used in the VEGAS and
 all the scripts used to analyze the data and generate the figures. The
 data also include supplementary data tables for the results of the DRGP2
 GWAS, MLM GWAS, and VEGAS.   
description: # Private QTLs underlie the genetic architecture of hierarchical size
 traits in *Drosophila* Dataset DOI:
 [10.5061/dryad.8sf7m0d36](https://doi.org/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 (µm^2^) #### 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
 (µm^2^) * 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).