Functional Study Of The Sugar Transporter Gene Family In The Genome-wide Range Of Neurospora Crassa

With the increase in the demand of bio-based fuels and chemicals,the expression of lignocellulase degrading enzymes and the rapid fermentation of biomass hydrolysates have become a hotspot.The sugar transportation is the first step in the use of lignocellulosic degradation products of cellulose fungi,which is not only the key component of fungal cells to absorb the vitro soluble sugar,but also plays an important role in the regulation of cellulase expression.Lignocellulose-degrading fungal Neurospora crassa has the ability to absorb and use a variety of monosaccharides and oligosaccharides.It is a model fungus that studies cellulose degradation,but more than half of the current prediction of sugar transporters are still unkown.In this study,Neurospora crassa was used as the research object to further identify the transporter substrat of filamentous fungal and explore the role of sugar transporters in the secretion of cellulase.In order to identify other unknown carbohydrate transporters in Neurospora crassa,we studied the substrate for the prediction sugar transporters from the whole genome level systematically and found that the two transporters(NCU01868 and NCU08152)have the function of transfer multiple hexose substrate.So they were named as NcHXT-1and NcHXT-2,respectively.The fluorescence resonance energy transfer technique(FRET)further confirmed that NcHXT-1/-2 had glucose-transporting function.NcHXT-1 /-2 was then overexpressed in EBY.VW4000,which was able to restore its ability to grow in the liquid medium of glucose,galactose or mannose and produce ethanol,indicating that both the genes have the potential application value of transforming the engineering strains to promote the use of biomass hydrolysates to synthesize bio-based chemicals.Transporter as a medium for the exchange of cells with the external environment,itplays roles in identifying the substrate and transmitting the molecular signal.It plays an important role in the induction of filamentous fungi cellulase.The results showed that NcHXT-2 could play a negative role in the expression of lignocellulase in Neurospora crassa.By construction of muti-gene mutant of NcHXT-2 and HGT-1/2,we found that NcHXT-2 had the redundancy with the high affinity glucose transporter HGT1/2.Although the deletion of NcHXT-2 resulted in up-regulation of expression of AMP cyclase gene,but the negative regulation of cellulase are also need to be confirmed.We found that NcHXT-1/-2 have homologous proteins in many cellulolytic fungi by phylogenetic tree analysis,which provided potential target for improving the enzyme activity in filamentous fungi.