## 2 Results
### TCDD Enriched for Mevalonate-Dependent Isoprenoid Biosynthesis
Abbreviations: 3-hydroxyl-3-methyl-clutaryl-CoA (HMG-CoA), (R)-5-Phosphomevalonate (mevalonate-5P), (R)-5-Diphosphomevalonate (mevalonate-5 PP), 2-C-Methyl-D-erythritol 4-phosphate (MEP), 4-(Cyltidine 5’-diphospho)-2-C-methyl-D-erythritol (CDP-ME), 4-(Cyltidine 5’-diphospho)-2-C-methyl-D-erythritol (DEP-ME-2P), 2-C-Methyl-D-erythritol 2,4-cyclodiphosphate (MEcPP), and 1-Hydroxy-2-methyl-2-butenyl 4-diphosphate (HMBPP). Vitamin K2 (Menaquinone) and Peptidoglycan Biosynthesis Pathways in Mouse NAFLD-Phenotypes and Gut Microbiology of Cirrhosis Patients  
In polyphenol diphosphate biosynthesis, IPP is recursively added to geranyl diphosphate (GPP) or farnesyl diphosphate (FPP) for polyphenol biosynthesis used in vitamin K2 (a.k.a., menaquinone) and peptidoglycan biosynthesis [41, 42]. TCDD enriched for heptaprenyl diphosphate synthase (EC 2.5.1.30) with major contributions from _[PERSON]. [PERSON]_ and _[PERSON]. [PERSON]_ (Figure 5). Figure 4: Mevalonate-dependent isoprenoid biosynthesis genes are enriched in a published metagenomics dataset of fecal samples from cirrhosis patients. Human3 analysis of fecal gut microbiomes in healthy (H, red, \(n=52\)), compensated (C, green, \(n=48\)), or decompensated (D, blue, \(n=44\)) cirrhosis for mevalonate-dependent and methyl-D-erythhtol 4-phosphate (MEP) pathways. Individual boxplots are numbered with the EC number matching the enzymatic step in pathway schematic. Significance is denoted with a red asterisk (*, adjusted \(p\)-values \(<0.05\)) compared to healthy group. Abbreviations. : 3-hydroxyl-3-methyl-clutaryl-CoA (HMG-CoA), (R)-5-Phosphomevalonate (mevalonate-5P), (R)-5-Diphosphomevalonate (mevalonate-5 PP), 2-C-Methyl-D-erythhtol 4-phosphate (MEP), 4-(C-Vtidine 5\({}^{\prime}\)-diphospho)-2-C-methyl-D-erythhtol (CDP-ME), 4-(C-Vtidine 5\({}^{\prime}\)-diphospho)-2-C-methyl-D-erythhtol (DEP-ME-2P), 2-C-Methyl-D-erythhtol 2,4-cyclodiphosphate (MEcPP), 1-Hydroxy-2-methyl-2-butenyl 4-diphosphate (HMBPP). As bacterial cell wall restructuring has been reported in response to bile acids and different levels of isoprenoid biosynthesis pathways were identified, peptidoglycan biosynthesis was also assessed [43]. Most genes encoding enzymes required for peptidoglycan biosynthesis were present in the metagenomic dataset (Figure 6A) with no changes observed following TCDD treatment. However, serine-type D-Ala-D-Ala carboxopeptidase (Figure 6a, EC 3.4.16.4, step 14), responsible for peptidoglycan polymer crosslinking, trended upwards. Additionally, most peptidoglycan biosynthesis EC numbers had annotations to _L. reuteri_ (Figure 6B). Overall, TCDD did not alter peptidoglycan synthesis related gene levels. De novo menaquinone biosynthesis requires chorismate and the addition of a polyprenol diphosphate (i.e., geranyl-geranyl diphosphate) (Figure 7A). Two alternative pathways exist for menaquinone biosynthesis, the o-succinylbenzoicate or futalosine route [44]. Only a few EC number annotations were detected for the futalosine pathway (EC 4.2.1.151 and EC 2.5.1.120), while all EC numbers were identified for the complete o-succinylbenzoicate menaquinone pathway (Figure 7A). Figure 5: Relative abundance of polyprenol transferase EC annotations identified in the mouse cecum metagenomic dataset. Stacked bar plots represent mean relative abundance of grouped EC numbers (\(n=3\)) and represent identified species that contributed to mean total abundance for each treatment group. The number of isoprenenyl diphosphate (IPP) and farnesyl diphosphate (FPP) molecules used for respective polyprenol biosynthesis are also denoted. Adjusted \(p\)-values were determined by the Maaslin2 R package. Abbreviations: isopentenyl diphosphate (IPP), geranyl diphosphate (GPP), polyprenyl diphosphate (polyphenyl-PP). Figure 6: Peptidoglycan biosynthesis was unchanged by TCDD. **(A)** Relative abundance of peptidoglycan biosynthesis EC numbers identified in the metagenomic dataset. **(B)** Relative abundance of only Lactobacillus species classified to peptidoglycan biosynthesis EC numbers.