3.5.4 Neural wiring
A range of genes and functional categories involved in neural wiring, including cell migration and adhesion, and neurite growth and synapse formation were affected by CO2 treatment, also mostly exhibiting an upregulation in the CNS (Table 3). There was small, coordinated upregulation of genes involved in cell migration functional categories, including ‘motor activity’, ‘actin binding’, ‘cell adhesion’ and ‘integrin complex’. Those genes positively correlated with CO2 treatment and all three activity traits in the CNS were enriched for similar functional categories: ‘actin filament binding’, ‘myosin complex’, ‘myofibril’, ‘motor activity’ and ‘ruffle membrane’. Genes with a role specifically in neuron migration and adhesion, and the related processes of dendrite and axon outgrowth and branching, dendritic spine formation, and synapse formation were also positively correlated with CO2 treatment and activity (rac1, ptpr, adgrb3, apbb1). One gene involved in neurite growth and branching (futsch) was negatively correlated with CO2 treatment and activity traits.
3.5.5 Transcription, RNA processing, and protein processing
There was generally an upregulation of genes and functional categories involved in transcription, RNA processing and protein processing in the CNS, and to a smaller extent in the eyes (Table 4). In the CNS, there was small, coordinated upregulation of genes belonging to functional categories involved in transcription, including ‘DNA duplex unwinding’, and its’ daughter term ‘3’-5’ DNA helicase activity’ was enriched in those genes positively correlated with CO2 treatment and all three activity traits (Figure 5). There was small, coordinated upregulation in the CNS of genes belonging to the functional category ‘nuclear pore’, and ‘nuclear pore’ was also enriched for those genes positively correlated with CO2 treatment and all three activity traits in the CNS (Figure 5). Genes involved in transcription were also DE; nme6, chrac1 and znf271 were upregulated in the CNS, and gtf2e2 downregulated in the eyes. There was small, coordinated upregulation of genes involved in ‘RNA processing’, ‘mRNA processing’ and ‘rRNA processing’, as well as functional categories involved in the processes and components of the spliceosome, which excises introns to produce mature mRNA (Figure 3). Components of the spliceosome (snrpa, snrnp200) were also identified as correlated with CO2 treatment and behaviours (Table 4).
A range of genes and functional categories involved in protein synthesis, folding and degradation/turnover were significantly upregulated in the CNS, and some were also upregulated in the eyes. Small, coordinated upregulation of genes involved in the initiation and process of translation as well as in functional categories for ribosomal components occurred in both tissues (Figure 3). Genes with similar functions were also positively correlated with COtreatment and activity traits in the CNS, including translation initiation factors (eif3b, eif3d), an elongation factor (eef1g), and ribosomal components (rpl23a, rpl4rpl7l, rps27a). Genes involved in protein translocation (stt3a, rpn1, rpn2, tmed2), and protein folding and quality control (pdia3, pdia4, pdia5, hspa5) were correlated with CO2 treatment and behaviours in the CNS. The endoplasmic reticulum associated degradation (ERAD) pathway involves protein ubiquitination followed by proteasomal degradation and we found a range of genes involved in this process to be affected by COtreatment. There was small, coordinated upregulation of genes in the ‘endoplasmic reticulum’ and five ubiquitin-related functional categories in the CNS, of ‘proteasome complex’ in both tissues, and of ‘peptide metabolic process’ in the eyes. Genes positively correlated with CO2 treatment and all three activity traits were enriched for ‘endoplasmic reticulum lumen’ and included genes coding for E3 ubiquitin ligases (cblb, ttc3) and a subunit for the 26S proteasome (rpn1). A gene coding for an E3 ubiquitin ligase (syvn1- b) was also upregulated in the CNS. Lysosomal degradation is another method for protein turnover, and a gene essential for lysosomal function (ykt6) was upregulated in both the CNS and eyes.