How To Use the Sparse Marginal Epistasis Test

Data Requirements and File Formats

SME is implemented in R and requires your genetic data to be in PLINK format, which consists of three files:

• .bim: Contains SNP information
• .bed: Contains genetic data
• .fam: Contains sample information

Note: sme() cannot handle missing genotypes. Prior to running sme(), use PLINK to remove variants with missing genotypes or impute them.

Additionally, your phenotype (trait) data should be in a separate file following PLINK’s phenotype format.

The sme() function includes parameters that let you control memory usage and computational resources. For detailed guidance on optimizing these settings for your system, please see our tutorial How To Optimize the Memory Requirements of SME.

Specifying SNPs for Analysis

When selecting which SNPs to analyze, you must provide their positions as 1-based indices. These indices correspond to the row numbers in your .bim file, where the first SNP is index 1, the second is index 2, and so on.

For complete details about all function parameters, please refer to the documentation of the sme() function.

# File inputs
plink_file <- "path/to/plink/file"
pheno_file <- "path/to/pheno/file"
mask_file <- "path/to/mask/file"

# Parameter inputs
chun_ksize <- 10
n_randvecs <- 10
n_blocks <- 10
n_threads <- 5

# 1-based Indices of SNPs to be analyzed
n_snps <- 100
snp_indices <- 1:n_snps

sme_result <- sme(
  plink_file,
  pheno_file,
  mask_file,
  snp_indices,
  chunk_size,
  n_randvecs,
  n_blocks,
  n_threads
)

This analysis uses simulated data for demonstration purposes. We simulated synthetic phenotypes from 5000 synthetic genotypes with 6000 SNPs. If you would like to learn how to simulate data, please refer to our tutorial How To Simulate Traits.

Understanding the Results

When you call the sme() function, it returns a list containing multiple elements. The most important one is called summary, which contains the main analysis results. These results are formatted as tidy data, making them compatible with popular R packages like ggplot2 and dplyr for further analysis and visualization.

Visualizing Genomic Associations

We use Manhattan plots to visualize genome-wide analyses because they effectively highlight strong associations between genetic variants and traits. In this case, the Manhattan plot specifically shows statistical epistasis (interactions between genes). For reference, we’ve marked the true causal SNPs (Single Nucleotide Polymorphisms) in green on the plot - these are the genetic variants we included in our simulation as having real effects.

sme_result$summary %>%
  ggplot(aes(
  x = index,
  y = -log10(p),
  color = true_gxg_snp
)) +
  geom_point() +
  xlab("Position") +
  labs(color = "Epistatic SNP")

Understanding Variance Components and Effect Sizes

SME estimates how much of the total trait variation can be explained by genetic interactions. In this simulation, we set the Phenotypic Variance Explained (PVE) to 5% for each SNP involved in epistatic interactions.

The plot below shows how well our method recovered these effects. It displays two distributions:

1. The estimated PVE for SNPs we know are truly involved in epistatic interactions
2. The estimated PVE for SNPs that have no real epistatic effects

The dashed line marks the true 5% PVE level we used in the simulation, allowing you to see how accurately the method estimated the actual effect sizes.

sme_result$summary %>%
  ggplot(aes(x = true_gxg_snp, y = pve, fill = true_gxg_snp)) +
  geom_boxplot() +
  geom_hline(yintercept = 0.05, color = "grey40", linetype = "dashed") +
  annotate("text", x = 0.8, y =  0.055,
           label = "True per SNP epistatic PVE", color = "black") +
  xlab("Epistatic SNP") +
  ylab("Phenotypic Variance Explained") +
  theme(legend.position = "none")

Narrow Sense Heritability Estimates

The SME method uses a linear mixed model to separate different sources of trait variation. One key component it estimates is narrow sense heritability (\(h^2\)), which measures how much trait variation can be explained by additive genetic effects.

The plot below breaks down the estimated sources of trait variation:

• “grm”: Shows narrow sense heritability (\(h^2\))
• “gxg”: Shows variance due to gene-by-gene interactions
• “error”: Shows unexplained variance, such as environmental effects

In our simulation, we set the true narrow sense heritability to 30%, shown as a dashed line in the plot. This reference line helps you evaluate how accurately SME estimated the genetic components of trait variation.

sme_result$vc_estimate %>%
  ggplot(aes(x = component, y = vc_estimate, fill = component)) +
  geom_boxplot() +
  geom_hline(yintercept = 0.3, color = "grey40", linetype = "dashed") +
  annotate("text", x = 0.7, y =  0.33,
           label = expression("True " * h^2), color = "black")  +
  xlab("Component") +
  ylab("Variance Component Estimate") +
  theme(legend.position = "none")

The estimate of the narrow-sense heritability \(h^2\) is much less variable because it is always informed by the same genetic relatedness matrix. In this small data example it overestimates the heritability but it is unbiased in general.