Gene sets that homogeneously respond to lrECM, regardless of the cell line, transformation status or spheroid morphology fell into 3 clusters: Cluster 7 (repressed) was highly enriched in mitochondrial and ribosomal functions, mRNA processing, and general metabolic processes, indicating the overall reduced growth, metabolic activity and proliferation of cells in 3D compared to monolayer culture.
Interestingly, subnetworks with conserved low connectivity were all metabolic processes indicating that these constitute separate modules in the networks.
We observed a pattern of changes in primary as well as secondary metabolic processes, indicating that our trait-deteriorated nematodes evolved massive metabolic changes.
The down-regulated gene lists revealed a significant reduction in metabolic processes, indicating that the animals were repartitioning their translation machinery away from normal housekeeping functions towards repair and regeneration (data not shown).
The high number of glycosylated proteins related to metabolic processes indicates the production of enough energy through the metabolism of carbohydrates and proteins for queen spawning and larval growth, which may be achieved by modulating the enzymatic efficiency [ 64].
However, the category with the second largest number of up-regulated sequences in the large Sept and Dec. fish was 'cellular process' and in the small Sept. and Dec. fish was 'metabolic process' indicating similarity in function across both size groups.
Further analysis of differentially expressed genes in the metabolic process category indicated that approximately 37% were related to nitrogen metabolism, 17% to phosphorus metabolism, and 15% to carbohydrate metabolism, together accounting for approximately 70% of genes in this category.
The network module 213 is enriched in the GO terms "nitrate reductase activity" and "nitrate reductase complex" whereas the network module 488 is enriched in the GO term "nitrogen compound metabolic process", which indicates that both networks may be involved in nitrogen metabolism.
Among the differentially expressed genes, a total of 719 sequences was categorized into 25 functional groups.Of these, the dominant terms were "catalytic activity", "binding", "cell", "cellular process" and "metabolic process", which indicated that these genes were enriched in the two transcriptome libraries.
Moreover, for groups associated with Al tolerance, the most enriched GO Slim terms are mitochondrion, transporter activity (Cytochrome b-c1 complex) and carbohydrates metabolic process (glyceraldehyde-3-phosphate dehydrogenase) indicating that miRNAs from these groups may mediate glycolysis and respiration process under Al stress conditions.
