scholarly journals Schixophyllum commune Aα Mating-Type Proteins, Y and Z, Form Complexes in All Combinations In Vitro

Genetics ◽  
1997 ◽  
Vol 147 (1) ◽  
pp. 117-123
Author(s):  
Yasuhiko Asada ◽  
Changli Yue ◽  
Jian Wu ◽  
Guang-Ping Shen ◽  
Charles P Novotny ◽  
...  
Keyword(s):  
Mating Type ◽  
Protein Interactions ◽  
Protein Complexes ◽  
Schizophyllum Commune ◽  
Mating Types ◽  
Partial Length ◽  
Z Protein ◽  
Specificity Domain

Abstract The Aα locus of the basidiomycete fungus, Schizophyllum commune, regulates sexual development via proteins Y and Z. Each Aα mating type encodes unique Y and Z isoforms. We used two isoforms of Y (Y4 and Y5) and two isoforms of Z (Z4 and Z5) in affinity assays of protein binding. These assays identified two types of protein interactions. Each full-length Y or Z protein binds to itself and other Y or Z proteins regardless of the Aα mating type from which they are encoded (i.e., mating-type independent binding). A second type of binding, detected with partial-length polypeptides, occurs only between N-terminal regions of Y and Z proteins encoded from different Aα mating types (e.g., Y4Z5 or Y5Z4); we refer to this binding as mating-type dependent binding. Deletion analysis shows that the Y4 specificity domain (an N-terminal region conferring recognition uniqueness to the Y4 isoform) is essential for mating-type dependent binding. Other regions of Y and Z are involved in mating-type independent binding. These results, obtained in vitro, raise the possibility that either of several protein complexes composed of Y and/or Z proteins may occur in vivo.

scholarly journals Interfacial Peptides as Affinity Modulating Agents of Protein-Protein Interactions

Biomolecules ◽  
2022 ◽  
Vol 12 (1) ◽  
pp. 106
Author(s):  
Pavel V. Ershov ◽  
Yuri V. Mezentsev ◽  
Alexis S. Ivanov
Keyword(s):  
Protein Interactions ◽  
Targeted Delivery ◽  
Protein Complexes ◽  
Protein Protein Interactions ◽  
Good Efficiency ◽  
Cellular Targets ◽  
Proteolytic Stability ◽  
Clinically Significant

The identification of disease-related protein-protein interactions (PPIs) creates objective conditions for their pharmacological modulation. The contact area (interfaces) of the vast majority of PPIs has some features, such as geometrical and biochemical complementarities, “hot spots”, as well as an extremely low mutation rate that give us key knowledge to influence these PPIs. Exogenous regulation of PPIs is aimed at both inhibiting the assembly and/or destabilization of protein complexes. Often, the design of such modulators is associated with some specific problems in targeted delivery, cell penetration and proteolytic stability, as well as selective binding to cellular targets. Recent progress in interfacial peptide design has been achieved in solving all these difficulties and has provided a good efficiency in preclinical models (in vitro and in vivo). The most promising peptide-containing therapeutic formulations are under investigation in clinical trials. In this review, we update the current state-of-the-art in the field of interfacial peptides as potent modulators of a number of disease-related PPIs. Over the past years, the scientific interest has been focused on following clinically significant heterodimeric PPIs MDM2/p53, PD-1/PD-L1, HIF/HIF, NRF2/KEAP1, RbAp48/MTA1, HSP90/CDC37, BIRC5/CRM1, BIRC5/XIAP, YAP/TAZ–TEAD, TWEAK/FN14, Bcl-2/Bax, YY1/AKT, CD40/CD40L and MINT2/APP.

scholarly journals Evolutionary diversification of protein–protein interactions by interface add-ons

2017 ◽  
Vol 114 (40) ◽  
pp. E8333-E8342 ◽  
Author(s):  
Maximilian G. Plach ◽  
Florian Semmelmann ◽  
Florian Busch ◽  
Markus Busch ◽  
Leonhard Heizinger ◽  
...  
Keyword(s):  
Protein Interaction ◽  
Protein Interactions ◽  
Protein Complexes ◽  
Evolutionary Strategy ◽  
Structural Level ◽  
Protein Protein Interactions ◽  
Protein Protein Interaction ◽  
Evolutionary Diversification

Cells contain a multitude of protein complexes whose subunits interact with high specificity. However, the number of different protein folds and interface geometries found in nature is limited. This raises the question of how protein–protein interaction specificity is achieved on the structural level and how the formation of nonphysiological complexes is avoided. Here, we describe structural elements called interface add-ons that fulfill this function and elucidate their role for the diversification of protein–protein interactions during evolution. We identified interface add-ons in 10% of a representative set of bacterial, heteromeric protein complexes. The importance of interface add-ons for protein–protein interaction specificity is demonstrated by an exemplary experimental characterization of over 30 cognate and hybrid glutamine amidotransferase complexes in combination with comprehensive genetic profiling and protein design. Moreover, growth experiments showed that the lack of interface add-ons can lead to physiologically harmful cross-talk between essential biosynthetic pathways. In sum, our complementary in silico, in vitro, and in vivo analysis argues that interface add-ons are a practical and widespread evolutionary strategy to prevent the formation of nonphysiological complexes by specializing protein–protein interactions.

scholarly journals Impact of Mating Type, Serotype, and Ploidy on the Virulence of Cryptococcus neoformans

2008 ◽  
Vol 76 (7) ◽  
pp. 2923-2938 ◽  
Author(s):  
Xiaorong Lin ◽  
Kirsten Nielsen ◽  
Sweta Patel ◽  
Joseph Heitman
Keyword(s):  
Cryptococcus Neoformans ◽  
Mating Type ◽  
Pathogenic Fungi ◽  
Mating Types ◽  
Type Locus ◽  
Virulence Potential ◽  
The Impact ◽  
Ad Hybrids

ABSTRACT Hybridization with polyploidization is a significant biological force driving evolution. The effect of combining two distinct genomes in one organism on the virulence potential of pathogenic fungi is not clear. Cryptococcus neoformans, the most common cause of fungal infection of the central nervous system, has a bipolar mating system with a and α mating types and occurs as A (haploid), D (haploid), and AD hybrid (mostly diploid) serotypes. Diploid AD hybrids are derived either from a-α mating or from unisexual mating between haploid cells. The precise contributions of increased ploidy, the effect of hybridization between serotypes A and D, and the combination of mating types to the virulence potential of AD hybrids have remained elusive. By using in vitro and in vivo characterization of laboratory-constructed isogenic diploids and AD hybrids with all possible mating type combinations in defined genetic backgrounds, we found that higher ploidy has a minor negative effect on virulence in a murine inhalation model of cryptococcosis. The presence of both mating types a and α in AD hybrids did not affect the virulence potential, irrespective of the serotype origin. Interestingly, AD hybrids with only one mating type behaved differently, with the virulence of αADα strains similar to that of other hybrids, while aADa hybrids displayed significantly lower virulence due to negative epistatic interactions between the Aa and Da alleles of the mating type locus. This study provides insights into the impact of ploidy, mating type, and serotype on virulence and the impact of hybridization on the fitness and virulence of a eukaryotic microbial pathogen.

scholarly journals Nucleosomes accelerate transcription factor dissociation

2013 ◽  
Vol 42 (5) ◽  
pp. 3017-3027 ◽  
Author(s):  
Yi Luo ◽  
Justin A. North ◽  
Sean D. Rose ◽  
Michael G. Poirier
Keyword(s):  
Single Molecule ◽  
Protein Interactions ◽  
Protein Complexes ◽  
High Specificity ◽  
Dissociation Rate ◽  
Duplex Dna ◽  
Recognition Sequence ◽  
Nucleosome Arrays

AbstractTranscription factors (TF) bind DNA-target sites within promoters to activate gene expression. TFs target their DNA-recognition sequences with high specificity by binding with resident times of up to hours in vitro. However, in vivo TFs can exchange on the order of seconds. The factors that regulate TF dynamics in vivo and increase dissociation rates by orders of magnitude are not known. We investigated TF binding and dissociation dynamics at their recognition sequence within duplex DNA, single nucleosomes and short nucleosome arrays with single molecule total internal reflection fluorescence (smTIRF) microscopy. We find that the rate of TF dissociation from its site within either nucleosomes or nucleosome arrays is increased by 1000-fold relative to duplex DNA. Our results suggest that TF binding within chromatin could be responsible for the dramatic increase in TF exchange in vivo. Furthermore, these studies demonstrate that nucleosomes regulate DNA–protein interactions not only by preventing DNA–protein binding but by dramatically increasing the dissociation rate of protein complexes from their DNA-binding sites.

scholarly journals The Specificity Determinant of the Y Mating-Type Proteins of Schizophyllum commune Is Also Essential for Y-Z Protein Binding

Genetics ◽  
1997 ◽  
Vol 145 (2) ◽  
pp. 253-260 ◽  
Author(s):  
Changli Yue ◽  
Michael Osier ◽  
Charles P Novotny ◽  
Robert C Ullrich
Keyword(s):  
Amino Acids ◽  
Protein Binding ◽  
Mating Type ◽  
Schizophyllum Commune ◽  
Mating Types ◽  
Mating Activity ◽  
Critical Residues ◽  
Mating Type Genes ◽  
Z Protein ◽  
Mating Tests

This paper concerns the manner in which combinatorial mating proteins of the fungus, Schizophyllum commune, recognize one another to form complexes that regulate target gene expression. In Schizophyllum, tightly linked Y and Z mating-type genes do not promote development in the combinations present in haploid strains (i.e., self combinations). When the Y and Z genes from two different mating types are brought together by the fusion of two haploid cells, the Y and Z proteins from different mating types recognize one another as nonself; form a complex and activate development. Several Y and Z alleles are present in the population and all nonself combinations of Y and Z alleles are equally functional. We have made chimeric genes among Y1, Y3, Y4 and Y5 and examined their mating-type specificities by transformation and mating tests. These studies show that the specificity of Y protein recognized by Z protein is encoded within a short region of N-terminal amino acids. The critical region is not precisely the same in each Y protein and in each Y-Z protein interaction. For Y3 protein compared with Y4 protein, the critical residues are in an N-terminal region of 56 amino acids (residues 17–72), with 40% identity and 65% similarity. Two-hybrid studies show that: the first 144 amino acids of Y4 protein are sufficient to bind Z3 and Z5 proteins, but not Z4 protein, and proteins deleted of the Y4 specificity region do not bind Z3, Z4 or Z5 protein. Thus the specificity determinant of the Y protein is essential for protein-protein recognition, Y-Z protein binding and mating activity.

scholarly journals Evidence for Interaction of Schizophyllum commune Y Mating-Type Proteins in Vivo

Genetics ◽  
2002 ◽  
Vol 160 (4) ◽  
pp. 1461-1467
Author(s):  
C Ian Robertson ◽  
Alexander McMahon Kende ◽  
Kurt Toenjes ◽  
Charles P Novotny ◽  
Robert C Ullrich
Keyword(s):  
Mating Type ◽  
Protein Interactions ◽  
Sexual Development ◽  
Fluorescent Protein ◽  
Schizophyllum Commune ◽  
Nuclear Localization Sequence ◽  
Type Locus ◽  
Localization Sequence ◽  
Green Fluorescent

Abstract The Aα mating-type locus of Schizophyllum commune regulates sexual development and contains the code for two proteins, Y and Z, which are thought to form a complex and function as a transcription factor. Import of these proteins into the nucleus may be an essential step in Aα-regulated sexual development. The Y proteins contain a bipartite basic sequence, which is an excellent candidate for a nuclear localization sequence (NLS), while Z proteins contain no such sequence. Here we describe experiments in which deletions were made in the putative NLS sequence of Y4. We show that this putative NLS is essential to the function of the Y protein and capable of mislocalizing green fluorescent protein (GFP) to the nucleus in Saccharomyces cerevisiae. Further, we describe genetic experiments that demonstrate the first Y-Y protein interactions in vivo. These results are consistent with our previously postulated hypothesis that the Y-Z complex is likely to be of a higher order than dimer.

scholarly journals Homeostatic Plasticity Requires Remodeling of the Homer-Shank Interactome

2020 ◽  
Author(s):  
Whitney E. Heavner ◽  
Haley Speed ◽  
Jonathan D. Lautz ◽  
Edward P. Gniffke ◽  
Karen B. Immendorf ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Cortical Neurons ◽  
Barrel Cortex ◽  
Protein Complexes ◽  
Sensory Deprivation ◽  
Interaction Network ◽  
Autism Spectrum ◽  
Homeostatic Plasticity

AbstractNeurons maintain constant levels of excitability using homeostatic scaling, which adjusts relative synaptic strength in response to large changes in overall activity. It is still unknown how homeostatic scaling affects network-level protein interactions in the synapse despite extensive reporting of individual scaling-associated transcriptomic and proteomic changes. Here, we assessed a glutamatergic synapse protein interaction network (PIN) composed of 380 binary interactions among 21 protein members to identify protein complexes altered by synaptic scaling in vitro and in vivo. In cultured cortical neurons, we observed widespread bidirectional PIN alterations during up- and downscaling that reflected rapid glutamate receptor shuttling via synaptic scaffold remodeling. Sensory deprivation of the barrel cortex caused a PIN response that reflected changes in mGluR tone and NMDAR-dependent metaplasticity, consistent with emerging models of homeostatic plasticity in the barrel cortex that restore excitatory/inhibitory balance. Mice lacking Homer1 or Shank3B did not undergo normal PIN rearrangements, suggesting that these Autism Spectrum Disorder (ASD)-linked proteins serve as structural hubs for synaptic homeostasis. Our approach demonstrates how changes in the protein content of synapses during homeostatic plasticity translate into functional PIN alterations that mediate changes in neuron excitability.

scholarly journals The Drosophila esc and E(z) Proteins Are Direct Partners in Polycomb Group-Mediated Repression

1998 ◽  
Vol 18 (5) ◽  
pp. 2825-2834 ◽  
Author(s):  
Clark A. Jones ◽  
Joyce Ng ◽  
Aidan J. Peterson ◽  
Kelly Morgan ◽  
Jeffrey Simon ◽  
...  
Keyword(s):  
Protein Complexes ◽  
Molecular Data ◽  
Polycomb Group ◽  
Physical Interaction ◽  
Binding Assays ◽  
Amino Acid Region ◽  
Z Protein ◽  
In Vitro Interaction

ABSTRACT The extra sex combs (esc) and Enhancer of zeste [E(z)] proteins are members of the Drosophila Polycomb group (Pc-G) of transcriptional repressors. Here we present evidence for direct physical interaction between the esc and E(z) proteins using yeast two-hybrid and in vitro binding assays. In addition, coimmunoprecipitation from embryo extracts demonstrates association of esc and E(z) in vivo. We have delimited the esc-binding domain of E(z) to an N-terminal 33-amino-acid region. Furthermore, we demonstrate that site-directed mutations in the esc protein previously shown to impair esc function in vivo disrupt esc-E(z) interactions in vitro. We also show an in vitro interaction between the heed and EZH1 proteins, which are human homologs of esc and E(z), respectively. These results suggest that the esc-E(z) molecular partnership has been conserved in evolution. Previous studies suggested that esc is primarily involved in the early stages of Pc-G-mediated silencing during embryogenesis. However, E(z) is continuously required in order to maintain chromosome binding by other Pc-G proteins. In light of these earlier observations and the molecular data presented here, we discuss how esc-E(z) protein complexes may contribute to transcriptional silencing by the Pc-G.

Photoinhibition in vivo and in vitro Involves Weakly Coupled Chlorophyll–Protein Complexes‡¶

2002 ◽  
Vol 75 (6) ◽  
pp. 613 ◽  
Author(s):  
Stefano Santabarbara ◽  
Ilaria Cazzalini ◽  
Andrea Rivadossi ◽  
Flavio M. Garlaschi ◽  
Giuseppe Zucchelli ◽  
...  
Keyword(s):  
Protein Complexes ◽  
Weakly Coupled ◽  
Chlorophyll Protein Complexes ◽  
Chlorophyll Protein

Probing the 14-3-3 Isoform-Specificity Profile of Interactions Stabilized by Fusicoccin A

2020 ◽  
Author(s):  
James Frederich ◽  
Ananya Sengupta ◽  
Josue Liriano ◽  
Ewa A. Bienkiewicz ◽  
Brian G. Miller
Keyword(s):  
Protein Interactions ◽  
Neurological Diseases ◽  
Adapter Proteins ◽  
Protein Protein Interactions ◽  
Specific Expression ◽  
Individual Client ◽  
Isoform Specificity ◽  
Fusicoccin A

Fusicoccin A (FC) is a fungal phytotoxin that stabilizes protein–protein interactions (PPIs) between 14-3-3 adapter proteins and their phosphoprotein interaction partners. In recent years, FC has emerged as an important chemical probe of human 14-3-3 PPIs implicated in cancer and neurological diseases. These previous studies have established the structural requirements for FC-induced stabilization of 14-3-3·client phosphoprotein complexes; however, the effect of different 14-3-3 isoforms on FC activity has not been systematically explored. This is a relevant question for the continued development of FC variants because there are seven distinct isoforms of 14-3-3 in humans. Despite their remarkable sequence and structural similarities, a growing body of experimental evidence supports both tissue-specific expression of 14-3-3 isoforms and isoform-specific functions <i>in vivo</i>. Herein, we report the isoform-specificity profile of FC <i>in vitro</i>using recombinant human 14-3-3 isoforms and a focused library of fluorescein-labeled hexaphosphopeptides mimicking the C-terminal 14-3-3 recognition domains of client phosphoproteins targeted by FC in cell culture. Our results reveal modest isoform preferences for individual client phospholigands and demonstrate that FC differentially stabilizes PPIs involving 14-3-3s. Together, these data provide strong motivation for the development of non-natural FC variants with enhanced selectivity for individual 14-3-3 isoforms.

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