PA-I and PA-II lectin interactions with the ABO(H) and P blood group glycosphingolipid antigens may contribute to the broad spectrum adherence ofPseudomonas aeruginosa to human tissues in secondary infections

Pseudomonas aeruginosa may cause serious infections in most human tissues/organs. Its adherence to them is mediated by a battery of adhesins including the PA-I and PA-II lectins, which are produced in this bacterium in high quantities. PA-I binds to thed-galactose of the erythrocyte glycosphingolipids exhibiting highest affinities for B and P^k (followed by P₁) antigens, while PA-II preferentially binds to thel-fucose of H, A and B antigens. IntactP. aeruginosa cells also exhibit a clear P^k and P₁ over p preference. Such affinities for the most common human ABH and P system antigens may underlie the widespread tissue infectivity and pathogenicity of this bacterium.     

Inferring primase-DNA specific recognition using a data driven approach

DNA–protein interactions play essential roles in all living cells. Understanding of how features embedded in the DNA sequence affect specific interactions with proteins is both challenging and important, since it may contribute to finding the means to regulate metabolic pathways involving DNA–protein interactions. Using a massive experimental benchmark dataset of binding scores for DNA sequences and a machine learning workflow, we describe the binding to DNA of T7 primase, as a model system for specific DNA–protein interactions. Effective binding of T7 primase to its specific DNA recognition sequences triggers the formation of RNA primers that serve as Okazaki fragment start sites during DNA replication.     

Scan2S: increasing the precision of PROSITE pattern motifs using secondary structure constraints

Sequence signature databases such as PROSITE, which include protein pattern motifs indicative of a protein's function, are widely used for function prediction studies, cellular localization annotation, and sequence classification. Correct annotation relies on high precision of the motifs. We present a new and general approach for increasing the precision of established protein pattern motifs by including secondary structure constraints (SSCs). We use Scan2S, the first sequence motif-scanning program to optionally include SSCs, to augment PROSITE pattern motifs. The constraints were derived from either the DSSP secondary structure assignment or the PSIPRED predictions for PROSITE-documented true positive hits. The secondary structure-augmented motifs were scanned against all SwissProt sequences, for which secondary structure predictions were precalculated. Against this dataset, motifs with PSIPRED-derived SSCs exhibited improved performance over motifs with DSSP-derived constraints. The precision of 763 of the 782 PSIPRED-augmented motifs remained unchanged or increased compared to the original motifs; 26 motifs showed an absolute precision increase of 10-30%. We provide the complete set of augmented motifs and the Scan2S program at http://physiology.med.cornell.edu/go/scan2s. Our results suggest a general protocol for increasing the precision of protein pattern detection via the inclusion of SSCs.     

Identification of GATC- and CCGG-recognizing Type II REases and their putative specificity-determining positions using Scan2S--a novel motif scan algorithm with optional secondary structure constraints

Restriction endonucleases (REases) are DNA-cleaving enzymes that have become indispensable tools in molecular biology. Type II REases are highly divergent in sequence despite their common structural core, function and, in some cases, common specificities towards DNA sequences. This makes it difficult to identify and classify them functionally based on sequence, and has hampered the efforts of specificity-engineering. Here, we define novel REase sequence motifs, which extend beyond the PD-(D/E)XK hallmark, and incorporate secondary structure information. The automated search using these motifs is carried out with a newly developed fast regular expression matching algorithm that accommodates long patterns with optional secondary structure constraints. Using this new tool, named Scan2S, motifs derived from REases with specificity towards GATC- and CGGG-containing DNA sequences successfully identify REases of the same specificity. Notably, some of these sequences are not identified by standard sequence detection tools. The new motifs highlight potential specificity-determining positions that do not fully overlap for the GATC- and the CCGG-recognizing REases and are candidates for specificity re-engineering.     

Primary ciliary dyskinesia caused by homozygous mutation in DNAL1, encoding dynein light chain 1

In primary ciliary dyskinesia (PCD), genetic defects affecting motility of cilia and flagella cause chronic destructive airway disease, randomization of left-right body asymmetry, and, frequently, male infertility. The most frequent defects involve outer and inner dynein arms (ODAs and IDAs) that are large multiprotein complexes responsible for cilia-beat generation and regulation, respectively. Although it has long been suspected that mutations in DNAL1 encoding the ODA light chain1 might cause PCD such mutations were not found. We demonstrate here that a homozygous point mutation in this gene is associated with PCD with absent or markedly shortened ODA. The mutation (NM_031427.3: c.449A>G; p.Asn150Ser) changes the Asn at position150, which is critical for the proper tight turn between the β strand and the α helix of the leucine-rich repeat in the hydrophobic face that connects to the dynein heavy chain. The mutation reduces the stability of the axonemal dynein light chain 1 and damages its interactions with dynein heavy chain and with tubulin. This study adds another important component to understanding the types of mutations that cause PCD and provides clinical information regarding a specific mutation in a gene not yet known to be associated with PCD.     

Effects of Low‐Dose Embryonic Thyroid Disruption and Rearing Temperature on the Development of the Eye and Retina in Zebrafish

Thyroid hormones are required for vertebrate development, and disruption of the thyroid system in developing embryos can result in a large range of morphologic and physiologic changes, including in the eye and retina. In this study, our anatomic analyses following low‐dose, chronic thyroid inhibition reveal that both methimazole (MMI) exposure and rearing temperature affect eye development in a time‐ and temperature‐dependent fashion. Maximal sensitivity to MMI for external eye development occurred at 65 hr postfertilization (hpf) for zebrafish reared at 28°C, and at 69 hpf for those reared at 31°C. Changes in eye diameter corresponded to changes in thickness of two inner retinal layers: the ganglion cell layer and the inner plexiform layer, with irreversible MMI‐induced decreases in layer thickness observed in larvae treated with MMI until 66 hpf at 28°C. We infer that maximal sensitivity to MMI between 65 and 66 hpf at 28°C indicates a critical period of thyroid‐dependent eye and retinal development. Furthermore, our results support previous work that shows spontaneous escape from MMI‐induced effects potentially due to embryonic compensatory actions, as our data show that embryos treated beyond the critical period generally resemble controls