Upstream open reading frames and Kozak regions of assembled transcriptome sequences from the spider Cupiennius salei. Selection or chance?

• We analyzed 5′ regions of a spider (Cupiennius salei) transcriptome.
• 81% of sequences had upstream open reading frames (uORFs).
• Kozak consensus regions were weaker for uORFs than main reading frames.
• mRNA abundances were negatively correlated with numbers of uORFs.
• Scrambled and random simulated 5′ region properties matched experimental data.

We assembled a new set of mRNA sequences from the leg hypodermis transcriptome of the wandering spider, Cupiennius salei. Each sequence was assembled to exhaustion in the 5′ direction to detect all upstream open reading frames (uORFs) both in-frame and out-of-frame with the main open reading frame (mORF). We also counted nucleotide probabilities before and after the START codon of the mORF to establish the optimum Kozak consensus sequence. More than 80% of 5′ sequences had uORFs before the mORF with a range of 1–16 uORFs. Kozak consensus strengths of uORFs were significantly weaker than mORFs. Random scrambling of 5′ nucleotide positions did not give significantly different numbers, sizes, or Kozak consensus strengths of uORFs. Random simulations of 5′ sequences using either equal or experimental distributions of nucleotides gave similar numbers of uORFs, with similar sizes and Kozak consensus strengths to experimental data. Abundance of mRNA for each gene was estimated by counting matching Illumina reads to assembled genes. Abundance was negatively correlated with numbers of uORFs, but not with 5′ length. Our data are compatible with a random model of 5′ mRNA sequence structure.