Functional Study of cAMP-Dependent Protein Kinase A in Penicillium oxalicum.

Signaling pathways play a crucial role in regulating cellulase production. The pathway mediated by signaling proteins plays a crucial role in understanding how cellulase expression is regulated. In this study, using affinity purification of ClrB, we have identified sixteen proteins that potentially interact with ClrB. One of the proteins, the catalytic subunit of cAMP-dependent protein kinase A (PoPKA-C), is an important component of the cAMP/PKA signaling pathway. Knocking out PoPKA-C resulted in significant decreases in the growth, glucose utilization, and cellulose hydrolysis ability of the mutant strain. Furthermore, the cellulase activity and gene transcription levels were significantly reduced in the ΔPoPKA-C mutant, while the expression activity of CreA, a transcriptional regulator of carbon metabolism repression, was notably increased. Additionally, deletion of PoPKA-C also led to earlier timing of conidia production. The expression levels of key transcription factor genes stuA and brlA, which are involved in the production of the conidia, showed significant enhancement in the ΔPoPKA-C mutant. These findings highlight the involvement of PoPKA-C in mycelial development, conidiation, and the regulation of cellulase expression. The functional analysis of PoPKA-C provides insights into the mechanism of the cAMP/PKA signaling pathway in cellulase expression in filamentous fungi and has significant implications for the development of high-yielding cellulase strains.

Phosducin-like protein PoPlp1 impacts cellulase and amylase expression and development in Penicillium oxalicum via the G protein-cAMP signaling pathway.

In this study, a phosducin-like protein, PoPlp1, was identified and functionally studied in the cellulase-producing strain Penicillium oxalicum 114-2. PoPlp1 was proven to participate in several biological processes, including mycelium development, conidiation, and expression of cellulases and amylases. With deletion of Poplp1, morphology and development varied significantly in ΔPoplp1. Colony growth, glucose utilization, and the hydrolysis capability of starch and cellulose were limited, whereas conidiation was enhanced. Based on detection of the levels of expression of transcription factors involved in asexual development, we conjectured that PoPlp1 is involved in conidiation via the major factor BrlA. We explored the effect of PoPlp1 on cellulase and amylase expression and observed that cellulase and amylase activity and major gene transcription levels were all dramatically reduced in ΔPoplp1. Deletion of PoPlp1 caused a decrease in intracellular cAMP levels, and the cellulase gene expression level of ΔPoplp1 was restored to a certain extent through external addition of cAMP. These findings demonstrate that PoPlp1 may affect cellulase and amylase expression by regulating cAMP concentration. To comprehensively explore the mechanism of PoPlp1 in regulating multiple biological processes, we performed a comparative transcriptomic analysis between strains P. oxalicum 114-2 and ΔPoplp1. The major cellulase and amylase genes were all downregulated, congrent with the results of real-time quantitative polymerase chain reaction analysis. The genes involved in the G protein-cAMP signaling pathway, including several G-protein-coupled receptors, one regulator of G protein signaling, and two cAMP phosphodiesterases, were disrupted by deletion of PoPlp1. These results confirm the positive function of PoPlp1 in the G protein-cAMP signaling pathway. This functional analysis of PoPlp1 will be very beneficial for further study of the regulatory mechanisms of cellulase expression and other biological processes in P. oxalicum 114-2 via the G protein-cAMP signaling pathway.

An actin-like protein PoARP9 involves in the regulation of development and cellulase and amylase expression in Penicillium oxalicum.

AIMS: In eukaryotic cells, chromatin remodelling complexes are essential for the accessibility of transcription factors to the specific regulating regions of downstream genes. Here, we identified an actin-like protein PoARP9 in cellulase production strain Penicillium oxalicum 114-2, which was an essential member of SWI/SNF complex. To investigate the physiological function of PoARP9 in transcriptional regulation, the coding gene Poarp9 was deleted in P. oxalicum 114-2. METHODS AND RESULTS: The absence of PoARP9 affected the colony growth on medium with glucose, cellulose or starch as sole carbon source. Meanwhile, the expression levels of major cellulase genes were all upregulated in ΔPoarp9 under the cellulase-inducing condition. In addition, the expression levels of amylase transcription activator AmyR as well as two major amylase genes were also increased in ΔPoarp9. CONCLUSIONS: These results demonstrated that chromatin remodelling affects the development and expression of cellulase and amylase in P. oxalicum. And the SWI/SNF complex member PoARP9 plays essential roles in these processes. SIGNIFICANCE AND IMPACT OF THE STUDY: This study provided new insights into the regulation of cellulase and development in P. oxalicum. And the regulatory function of SWI/SNF complex member ARP9 towards cellulase and amylase expression in P. oxalicum was verified for the first time.

A Highly Efficient Fluorescent Sensor Based on AIEgen for Detection of Nitrophenolic Explosives.

The detection of nitrophenolic explosives is important in counterterrorism and environmental protection, but it is still a challenge to identify the nitroaromatic compounds among those with a similar structure. Herein, a simple tetraphenylethene (TPE) derivative with aggregation-induced emission (AIE) characteristics was synthesized and used as a fluorescent sensor for the detection of nitrophenolic explosives (2, 4, 6-trinitrophenol, TNP and 2, 4-dinitrophenol, DNP) in water solution and in a solid state with a high selectivity. Meanwhile, it was found that only hydroxyl containing nitrophenolic explosives caused obvious fluorescence quenching. The sensing mechanism was investigated by using fluorescence titration and 1H NMR spectra. This simple AIE-active probe can potentially be applied to the construction of portable detection devices for explosives.

A Homeodomain-Containing Transcriptional Factor PoHtf1 Regulated the Development and Cellulase Expression in Penicillium oxalicum.

Homeodomain-containing transcription factors (Htfs) play important roles in animals, fungi, and plants during some developmental processes. Here, a homeodomain-containing transcription factor PoHtf1 was functionally characterized in the cellulase-producing fungi Penicillium oxalicum 114-2. PoHtf1 was shown to participate in colony growth and conidiation through regulating the expression of its downstream transcription factor BrlA, the key regulator of conidiation in P. oxalicum 114-2. Additionally, PoHtf1 inhibited the expression of the major cellulase genes by coordinated regulation of cellulolytic regulators CreA, AmyR, ClrB, and XlnR. Furthermore, transcriptome analysis showed that PoHtf1 participated in the secondary metabolism including the pathway synthesizing conidial yellow pigment. These data show that PoHtf1 mediates the complex transcriptional-regulatory network cascade between developmental processes and cellulolytic gene expression in P. oxalicum 114-2. Our results should assist the development of strategies for the metabolic engineering of mutants for applications in the enzymatic hydrolysis for biochemical production.

The effect of two different biochars on remediation of Cd-contaminated soil and Cd uptake by Lolium perenne.

Biochar can be widely used to reduce the bioavailability of heavy metals in contaminated soil because of its adsorption capacity. But there are few studies about the effects of biochar on cadmium uptake by plants in soil contaminated with cadmium (Cd). Therefore, an incubation experiment was used to investigate the effects of rice straw biochar (RSBC) and coconut shell biochar (CSBC) on Cd immobilization in contaminated soil and, subsequently, Cd uptake by Lolium perenne. The results showed that the microbial counts and soil enzyme activities were significantly increased by biochar in Cd-contaminated soil, which were consistent with the decrease of the bioavailability of Cd by biochar. HOAc-extractable Cd in soil decreased by 11.3-22.6% in treatments with 5% RSBC and by 7.2-17.1% in treatments with 5% CSBC, respectively, compared to controls. The content of available Cd in biochar treatments was significantly lower than in controls, and these differences were more obvious in treatment groups with 5% biochar. The Cd concentration in L. perenne reduced by 4.47-26.13% with biochar. However, the biomass of L. perenne increased by 1.35-2.38 times after adding biochar amendments. So, Cd uptake by whole L. perenne was augmented by RSBC and CSBC. Accordingly, this work suggests that RSBC and CSBC have the potential to be used as a useful aided phytoremediation technology in Cd-contaminated soil.

Alleviating the toxicity of heavy metals by combined amendments in cultivated bag of Pleurotus cornucopiae.

The substrate of mushroom can be polluted with heavy metals and subsequently contaminate mushroom, which requires alternative solutions to reduce associated environmental and human health risks. The effects of amendment application on alleviating Cu and Cd toxicities to Pleurotus cornucopiae were investigated in a cultivated bag experiment conducted with the naturally contaminated substrate. Addition of combined amendments (sodium bentonite, silicon fertilizer, activated carbon, and potassium dihydrogen phosphate) increased the P. cornucopiae biomass and substrate pH. Cu and Cd concentration in P. cornucopiae as well as the available Cu and Cd in substrate reduced for the presence of amendments, and the silicon fertilizer had the biggest inhibition on metal uptake. The smallest amount of Cu and Cd in P. cornucopiae was only 30.8 and 5.51% of control, respectively. Moreover, application of amendments also decreased malondialdehyde (MDA) and hydrogen peroxide (H2O2) level in metal-stressed mushroom by 4.38-53.74 and 8.90-58.42% relative to control, respectively. The decreased oxidative stress could well contribute to the growth of P. cornucopiae, and the elevated substrate pH might lead to the lower metal availability, thus resulting in the reduction of metal accumulation in mushroom. These above results suggest that application of combined amendments in mushroom substrate could be implemented in a general scheme aiming at controlling metal content in P. cornucopiae.