NMR structure of the human Rad18 zinc finger in complex with ubiquitin defines a class of UBZ domains in proteins linked to the DNA damage response.

Ubiquitin-mediated interactions are critical for the cellular DNA damage response (DDR). Therefore, many DDR-related proteins contain ubiquitin-binding domains, including ubiquitin-binding zinc fingers (UBZs). The majority of these UBZ domains belong to the C2H2 (type 3 Polη-like) or C2HC (type 4 Rad18-like) family. We have used nuclear magnetic resonance (NMR) spectroscopy to characterize the binding to ubiquitin and determine the structure of the type 4 UBZ domain (UBZ4) from human Rad18, which is a key ubiquitin ligase in the DNA damage tolerance pathway responsible for monoubiquitination of the DNA sliding clamp PCNA. The Rad18-UBZ domain binds ubiquitin with micromolar affinity and adopts a ß1-ß2-α fold similar to the previously characterized type 3 UBZ domain (UBZ3) from the translesion synthesis DNA polymerase Polη. However, despite nearly identical structures, a disparity in the location of binding-induced NMR chemical shift perturbations shows that the Rad18-UBZ4 and Polη-UBZ3 domains bind ubiquitin in distinctly different modes. The Rad18-UBZ4 domain interacts with ubiquitin with the α-helix and strand ß1 as shown by the structure of the Rad18-UBZ domain-ubiquitin complex determined in this work, while the Polη-UBZ3 domain exclusively utilizes the α-helix. Our findings suggest the existence of two classes of UBZ domains in DDR-related proteins with similar structures but unique ubiquitin binding properties and provide context for further study to establish the differential roles of these domains in the complex cellular response to DNA damage.

Persistent delay in maturation of the developing gut microbiota in infants with cystic fibrosis.

The healthy human infant gut microbiome undergoes stereotypical changes in taxonomic composition between birth and maturation to an adult-like stable state. During this time, extensive communication between microbiota and the host immune system contributes to health status later in life. Although there are many reported associations between microbiota compositional alterations and disease in adults, less is known about how microbiome development is altered in pediatric diseases. One pediatric disease linked to altered gut microbiota composition is cystic fibrosis (CF), a multi-organ genetic disease involving impaired chloride secretion across epithelia and heightened inflammation both in the gut and at other body sites. Here, we use shotgun metagenomics to profile the strain-level composition and developmental dynamics of the infant fecal microbiota from several CF and non-CF longitudinal cohorts spanning from birth to greater than 36 months of life. We identify a set of keystone species whose prevalence and abundance reproducibly define microbiota development in early life in non-CF infants, but are missing or decreased in relative abundance in infants with CF. The consequences of these CF-specific differences in gut microbiota composition and dynamics are a delayed pattern of microbiota maturation, persistent entrenchment in a transitional developmental phase, and subsequent failure to attain an adult-like stable microbiota. We also detect the increased relative abundance of oral-derived bacteria and higher levels of fungi in CF, features that are associated with decreased gut bacterial density in inflammatory bowel diseases. Our results define key differences in the gut microbiota during ontogeny in CF and suggest the potential for directed therapies to overcome developmental delays in microbiota maturation.

Bisphenol A promotes stress granule assembly and modulates the integrated stress response.

Bisphenol-A (BPA) is a ubiquitous precursor of polycarbonate plastics that is found in the blood and serum of >92% of Americans. While BPA has been well documented to act as a weak estrogen receptor (ER) agonist, its effects on cellular stress are unclear. Here, we demonstrate that high-dose BPA causes stress granules (SGs) in human cells. A common estrogen derivative, ß-estradiol, does not trigger SGs, indicating the mechanism of SG induction is not via the ER pathway. We also tested other structurally related environmental contaminants including the common BPA substitutes BPS and BPF, the industrial chemical 4-nonylphenol (4-NP) and structurally related compounds 4-EP and 4-VP, as well as the pesticide 2,4-dichlorophenoxyacetic acid (2,4-D). The variable results from these related compounds suggest that structural homology is not a reliable predictor of the capacity of a compound to cause SGs. Also, we demonstrate that BPA acts primarily through the PERK pathway to generate canonical SGs. Finally, we show that chronic exposure to a low physiologically relevant dose of BPA suppresses SG assembly upon subsequent acute stress. Interestingly, this SG inhibition does not affect phosphorylation of eIF2α or translation inhibition, thus uncoupling the physical assembly of SGs from translational control. Our work identifies additional effects of BPA beyond endocrine disruption that may have consequences for human health.