Biology subjects

Altegoer, F.

Publications and source records attributed to Altegoer, F..

4 recordsLinked to original sources

bioRxiv2022

Structural and functional analysis of the cerato-platanin-like effector protein Cpl1 suggests diverging functions in smut fungi

Plant pathogenic fungi are causative agents of the majority of plant diseases and can lead to severe crop loss in infected populations. Fungal colonization is achieved by combining different strategies, such as avoiding and counteracting the plant immune system and manipulating the host metabolome. Of major importance are effector proteins secreted by the fungi that fulfill diverse functions to support the infection process. Most of these proteins are highly specialized and structural and biochemical information is often absent. Here, we present the atomic structures of the cerato-platanin-like protein Cpl1 from Ustilago maydis and its homolog Uvi2 from Ustilago hordei. Both proteins adopt a double-{Psi}-{beta}-barrel architecture reminiscent of cerato-platanin proteins, a class so far not described in smut fungi. Our structure-function analysis shows that Cpl1 binds to soluble chitin fragments via two extended grooves at the dimer interface of the two monomer molecules. This carbohydrate-binding mode has not been observed previously and expands the repertoire of chitin-binding proteins. Cpl1 localizes to the cell wall of U. maydis and specifically enriches cell-wall degrading and -decorating proteins during maize infection. The architecture of Cpl1 harboring four surface exposed loop regions supports the idea that it might play a role in spatial coordination of these proteins. While deletion of cpl1 has only mild effects on the virulence of U. maydis, a recent study showed that deletion of uvi2 strongly impairs U. hordei virulence. Our structural comparison between Cpl1 and Uvi2 reveals sequence variations in the loop regions which might explain a diverging function.

biochemistry↗

bioRxiv2021

Structure and mechanistic features of the prokaryotic minimal RNase P

Endonucleolytic removal of 5-leader sequences from tRNA precursor transcripts (pre-tRNAs) by RNase P is essential for protein synthesis. Beyond RNA-based RNase P enzymes, protein-only versions of the enzyme exert this function in various Eukarya (there termed PRORPs) and in some bacteria (Aquifex aeolicus and close relatives); both enzyme types belong to distinct subgroups of the PIN domain metallonuclease superfamily. Homologs of Aquifex RNase P (HARPs) are also expressed in some other bacteria and many archaea, where they coexist with RNA-based RNase P and do not represent the main RNase P activity. Here we solved the structure of the bacterial HARP from Halorhodospira halophila by cryo-EM revealing a novel screw-like dodecameric assembly. Biochemical experiments demonstrate that oligomerization is required for RNase P activity of HARPs. We propose that the tRNA substrate binds to an extended spike-helix (SH) domain that protrudes from the screw-like assembly to position the 5-end in close proximity to the active site of the neighboring dimer subunit. The structure suggests that eukaryotic PRORPs and prokaryotic HARPs recognize the same structural elements of pre-tRNAs (tRNA elbow region and cleavage site). Our analysis thus delivers the structural and mechanistic basis for pre-tRNA processing by the prokaryotic HARP system.

biochemistry↗

bioRxiv2021

The CTPase activity of ParB acts as a timing mechanism to control the dynamics and function of prokaryotic DNA partition complexes

DNA partitioning CTPases of the ParB family mediate the segregation of bacterial chromosomes and low-copy number plasmids. They act as DNA-sliding clamps that are loaded at parS motifs in the centromeric region of target DNA molecules and then spread laterally to form large nucleoprotein complexes that serve as docking points for the DNA segregation machinery. Here, we identify conformational changes that underlie the CTP- and parS-dependent closure of ParB clamps. Moreover, we solve crystal structures of ParB in the pre- and post-hydrolysis state and provide insights into the catalytic mechanism underlying nucleotide hydrolysis. The characterization of CTPase-deficient ParB variants reveals that CTP hydrolysis serves as a timing mechanism to control the sliding time of ParB. Hyperstable clamps are trapped on the DNA, leading to excessing spreading and severe chromosome segregation defects in vivo. These findings clarify the role of the ParB CTPase cycle in partition complex dynamics and function and thus complete our understanding of this prototypic CTP-dependent molecular switch.

microbiology↗

bioRxiv2021

Identification and characterization of two transmembrane proteins required for virulence of Ustilago maydis

Smut fungi comprise a large group of biotrophic phytopathogens infecting important crops such as wheat and corn. Through the secretion of effector proteins, the fungus actively suppresses plant immune reactions and modulates its hosts metabolism. Consequently, how soluble effector proteins contribute to virulence is already characterized in a range of phytopathogens. However, membrane-associated virulence factors have been much less studied to date. Here, we investigated six transmembrane (TM) proteins that show elevated gene expression during biotrophic development of the maize pathogen Ustilago maydis. We show that two of the six proteins, named Vmp1 and Vmp2 (virulence-associated membrane protein), are essential for the full virulence of U. maydis. The deletion of the corresponding genes lead to a substantial attenuation in the virulence of U. maydis. Furthermore, both are conserved in various related smuts and contain no domains of known function. Our biochemical analysis clearly shows that Vmp1 and Vmp2 are membrane-associated proteins, potentially localizing to the U. maydis plasma membrane. Mass photometry and light scattering suggest that Vmp1 mainly occurs as a monomer, while Vmp2 is dimeric. Notably, the large and partially unstructured C-terminal domain of Vmp2 is crucial for virulence while not contributing to dimerization. Taken together, we here provide an initial characterization of two membrane proteins as virulence factors of U. maydis.

plant biology↗