An improved characterization of horse ( Equus caballus

Abstract Peripheral blood lymphocytes were cultured and treated for early- and late-BrdU incorporation to perform replicating G- and R-banding patterns, respectively. Slides were treated for GBG-, RBA- and RBG-banding techniques. Improved banded karyotypes at early- (350 bands) and pro-metaphase (500 bands) stage were performed and GBG- and RBA-banded prometaphase karyotypes were presented for the first time on this species. All chromosomes, including the small acrocentrics, show clear and distinguishable G- and R-banding patterns. Chromosome identification followed the latest chromosome standard nomenclature (ISCNH 1997). This study is also our contribution to further standard karyotype attempts at the prometaphase stage

Functional differences between E. Coli and eskape pathogen groes/groel

As the GroES/GroEL chaperonin system is the only bacterial chaperone that is essential under all conditions, we have been interested in the development of GroES/GroEL inhibitors as potential antibiotics. Using Escherichia coli GroES/GroEL as a surrogate, we have discovered several classes of GroES/GroEL inhibitors that show potent antibacterial activity against both Gram-positive and Gram-negative bacteria. However, it remains unknown if E. coli GroES/GroEL is functionally identical to other GroES/GroEL chaperonins and hence if our inhibitors will function against other chap-eronins. Herein we report our initial efforts to characterize the GroES/GroEL chapero-nins from clinically significant ESKAPE pathogens (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species). We used complementation experiments in GroES/GroEL-deficient and-null E. coli strains to report on exogenous ESKAPE chaperone function. In GroES/ GroEL-deficient (but not knocked-out) E. coli, we found that only a subset of the ESKAPE GroES/GroEL chaperone systems could complement to produce a viable orga-nism. Surprisingly, GroES/GroEL chaperone systems from two of the ESKAPE pathogens were found to complement in E. coli, but only in the strict absence of either E. coli GroEL (P. aeruginosa) orbothE. coli GroES and GroEL (E. faecium). In addition, GroES/ GroEL from S. aureus was unable to complement E. coli GroES/GroEL under all condi-tions. The resulting viable strains, in which E. coli groESL was replaced with ESKAPE groESL, demonstrated similar growth kinetics to wild-type E. coli, butdisplayedan elongated phenotype (potentially indicating compromised GroEL function) at some temperatures. These results suggest functional differences between GroES/GroEL chaperonins despite high conservation of amino acid identity. IMPORTANCE The GroES/GroEL chaperonin from E. coli has long served as the model system for other chaperonins. This assumption seemed valid because of the high conservation between the chaperonins. It was, therefore, shocking to discover ESKAPE pathogen GroES/GroEL formed mixed-complex chaperonins in the presence of E. coli GroES/GroEL, leading to loss of organism viability in some cases. Complete replacement of E. coli groESL with ESKAPE groESL restored organism viability, but produced an elongated phenotype, suggesting differences in chaperonin function, including client specificity and/or refolding cycle rates. These data offer important mechanistic insight into these remarkable machines, and the new strains developed allow for the synthesis of homogeneous chaperonins for biochemical studies and to further our efforts to de-velop chaperonin-targeted antibiotics. © 2021 Sivinski et al.Open access journalThis item from the UA Faculty Publications collection is made available by the University of Arizona with support from the University of Arizona Libraries. If you have questions, please contact us at [email protected]