Transcriptional upregulation of four genes of the lysine biosynthetic pathway by homocitrate accumulation in Penicillium chrysogenum: homocitrate as a sensor of lysine-pathway distress

Abstract

The lysine biosynthetic pathway has to supply large amounts of α-aminoadipic acid for penicillin biosynthesis in Penicillium chrysogenum. In this study, we have characterized the P. chrysogenum L2 mutant, a lysine auxotroph that shows highly increased expression of several lysine biosynthesis genes (lys1, lys2, lys3, lys7). The L2 mutant was found to be deficient in homoaconitase activity since it was complemented by the Aspergillus nidulans lysF gene. We have cloned a gene (named lys3) that complements the L2 mutation by transformation with a P. chrysogenum genomic library, constructed in an autonomous replicating plasmid. The lys3-encoded protein showed high identity to homoaconitases. In addition, we cloned the mutant lys3 allele from the L2 strain that showed a G1534 to A1534 point mutation resulting in a Gly495 to Asp495 substitution. This mutation is located in a highly conserved region adjacent to two of the three cysteine residues that act as ligands to bind the iron–sulfur cluster required for homoaconitase activity. The L2 mutant accumulates homocitrate. Deletion of the lys1 gene (homocitrate synthase) in the L2 strain prevented homocitrate accumulation and reverted expression levels of the four lysine biosynthesis genes tested to those of the parental prototrophic strain. Homocitrate accumulation seems to act as a sensor of lysine-pathway distress, triggering overexpression of four of the lysine biosynthesis genes.

The AMP-Activated Protein Kinase (AMPK) Positively Regulates Lysine Biosynthesis Induced by Citric Acid in Flammulina filiformis

“…This work found that lysine content increased by nearly 13% under optimal citric acid induction conditions, with the gene expression levels of hcs , hac, and hah being 2.67, 1.97, and 1.90 times higher than the control, respectively. Citric acid induction creates more metabolic flux into lysine biosynthesis, along with upregulating lysine-biosynthesis-related enzymes, and thus significantly increases the lysine content in F. filiformis , which is in line with previous studies proving that synthetic substrate accumulation accelerates biosynthesis pathways [ 30 , 32 ].…”

“…In S. cerevisiae , the α-aminoadipate pathway is initiated by homocitrate synthase, which catalyzes the first and rate-limiting step in the pathway and is highly regulated to conserve the use of resources [ 17 ]. Teves et al found that homocitrate was a regulatory substance controlling the lysine biosynthesis pathway by mutating the homoaconitase of Penicillium chrysogenum to achieve a significant accumulation of intracellular homocitrate, which caused a significant increase in the expression of lysine-biosynthesis-related genes, whereas the genes were normally expressed in a lysine auxotroph defective in the homocitrate synthase or in another strain disrupted in the α-aminoadipate reductase [ 30 ]. O’Doherty et al exposed a saccharopine dehydrogenase gene mutant (lys1Δ) strain of S. cerevisiae to lipid oxidant-linoleic acid hydroperoxide (LoaOOH) using a medium containing sufficient lysine, finding that it grew significantly slower compared to the wild-type BY4743 strain [ 31 ].…”

The fungal α‐aminoadipate pathway for lysine biosynthesis requires two enzymes of the aconitase family for the isomerization of homocitrate to homoisocitrate

“…However, whether this homoaconitase performs both reactions that are required for the isomerization has not been convincingly shown. Investigations on homoaconitase mutants from the filamentous fungi Aspergillus nidulans and Penicillium chrysogenum revealed an accumulation of homocitrate, but not homoaconitate (Weidner et al ., ; Teves et al ., ). Thus, it was assumed that homoaconitases from filamentous fungi catalyse both reactions.…”

TrLys9 participates in fungal development and lysine biosynthesis in <i>Trichoderma reesei</i>

“…A lysine biosynthesis branching intermediate α-aminoadipic acid is also one of the the precursors of the δ-L-α-aminoadipyl-L-cysteinyl-D-valine (ACV) tripeptide in the synthesis of penicillin (Esmahan et al, 1994). Indeed, one study found that penicillin production was impaired in P. chrysogenum mutants in which the branching point of the lysine biosynthesis pathway was blocked due to unavailability of α-aminoadipic acid (Teves et al, 2009), while another found that disruption of the lys2 gene led to penicillin overproduction due to an increased pool of α-aminoadipic acid (Casqueiro et al,. 1999).…”