Analysis of Proteome Dynamics in Early-Stage Sporulation-Inhibited Variants of Parageobacillus thermoglucosidasius

Parageobacillus thermoglucosidasius is a thermophilic Gram-positive that is emerging as a platform for bioproduction. However, one potential limitation of P. thermoglucosidasius is its natural tendency to sporulate. As a non-model organism, the complex regulatory system that governs sporulation in P. thermoglucosidasius remains poorly characterised. To advance current understanding, this study presents a comparative characterisation and proteomic analysis of the P. thermoglucosidasius wildtype strain alongside four early-stage sporulation-inhibited variants. To inhibit sporulation, the genes spo0A, spo0B, spo0F, and sigF were targeted for deletion, as based on their crucial regulatory roles in the sporulation pathway of Bacillus subtilis. Microscopic analysis indicated that the Δspo0A, Δspo0F, and ΔsigF strains were sporulation-suppressed while the Δspo0B strain sporulated at low levels. Proteomics data were obtained from four different growth stages and the resulting expression profiles were compared. Consistent with the regulatory network of B. subtilis, the Δspo0A, Δspo0B, and Δspo0F strains exhibited largely inactive sporulation pathways, while the ΔsigF strain retained some early regulatory functions. Additionally, two co-expression modules comprising approximately 300 genes were identified and linked to the P. thermoglucosidasius sporulation pathway. Overall, these results expand the understanding of the sporulation network of P. thermoglucosidasius and provide a foundation for future engineering of non-sporulating variants.