Sequence-specific 'H and '-N resonance asignments have been determined for the major cold shock protein (CspA) from Escherichia coli with recently developed three-dimensional triple-resonance NMR experiments. By use of these asgments, five antp lel 1-strands were identified from analysis of NMR data. Strands 1-4 have a clsical 3-2-1-4 Greek key -sheet topology and there are two 1-bulges, at positions Lys'0-Trp" and Gly"5-Asn". Threedimensional structures of CspA were generated from NMR data by using smulated annling with molecular dynamics. The overall chain fold of CspA Is a 1-barrel structure, with a tightly packed hydrophobic core. Two-dimensional isotopeedited pulsed-field gradient 15N-1H heteronuclear singlequantum coherence spectroscopy was used to chracerie the "N-1H fingerprint spectrum with and without a 24-base oligodeoxyribonucleotide, 5'-AACGGTTTGACGTACAGAC-CATTA-3'. Protein-DNA complex formation perturbs a subset of the amide resonances that are located mostly on one face of the CspA molecule. This portion of the CspA molecular surface includes two putative RNA-binding sequence motifs which contribute to an unusual cluster of eight surface aromatic side chais: Trpol Phel", Phe", P , Phe3, His33, Phe3, and Tyr42. These surface aromatic groups, and also residues Lys"6, Ser", and Lys"' located on this same face of CspA, are higly conserved in the family of CspA homologues. These isotopeedited pulsed-field gradient NMR data provide a low-resolution mapping of a DNA-binding epitope on CspA.eukaryotic Y-box family of transcription-regulating proteins (6)(7)(8) (4,18), and these also contain RNP1 and RNP2 sequence motifs. The three-dimensional structure of one of these, the major cold shock protein CspB from Bacillus subtilis, has been determined recently by solution NMR (19) and x-ray crystallography (16). CspB has structural homology with several nucleic acid-binding proteins and forms complexes with ssDNA oligomers containing a CCAAT site in gel retardation experiments (16,19). In this paper we describe the overall chain fold of E. coli CspA determined in solution by NMR spectroscopy and demonstrate that CspA forms a complex with a 24-base oligodeoxyribonucleotide corresponding in sequence to the 5' leader region of cspA mRNA. 11 Many bacteria respond to low temperature stress by production ofcold shock proteins (1-5). In Escherichia coli this cold shock response involves enhanced production of at least 14 proteins (1, 2). One of these, the major cold shock protein (CspA), is not detectable when cells are grown at 370C but is produced at levels of 10-15% of total protein synthesis when cells are shifted to 10-15TC (2), with expression peaking 60-120 min following cold shock. The gene for CspA (cspA) has been cloned and sequenced (2), and primer extension studies indicate that levels of cspA transcripts also increase and then decrease in the cold shock response, paralleling the production kinetics of CspA protein. The transient nature of cold shock-induced cspA expression indicates a regulator...

Background Keratinocytes at wound margins undergo partial epithelial to mesenchymal transition (EMT). Based on previous in vitro and ex vivo findings, Slug (Snai2), a transcriptional regulator of EMT in development, may play an important role in this process. Objectives This study was designed to validate an in vivo role for Slug in wound healing. Methods Excisional wounds in Slug null and wild type mice were examined histologically at 6, 24, 48, and 72 h after wounding; reepithelialization was measured and immunohistochemistry for keratins 8, 10, 14, and 6 and E-cadherin was performed. In 20 Slug null and 20 wild type mice exposed three times weekly to two minimal erythemal doses of UVR, the development of non-healing cutaneous ulcers was documented. Ulcers were examined histologically and by immunohistochemistry. Results The reepithelialization component of excisional wound healing was reduced 1.7-fold and expression of the Slug target genes keratin 8 and E-cadherin was increased at wound margins in Slug null compared to wild type mice. In contrast, no differences in expression of keratins 10 or 14 or in markers of proliferation K6 and Ki-67 were observed. Forty per cent of Slug null mice but no wild type mice developed non-healing cutaneous ulcers in response to chronic UVR. Keratinocytes at ulcer margins expressed high levels of keratin 8 and retained E-cadherin expression, thus resembling excisional wounds. Conclusion Slug is an important modulator of successful wound repair in adult tissue and may be critical for maintaining epidermal integrity in response to chronic injury.

Abstract. Submissions to the University of Tennessee pathology service from June 1999 to June 2008 were searched for feline cases of tumors involving the eyelids or nictitans. Forty-three tumors were identified. The average age at diagnosis was 10.4 years. Significantly more males than females had eyelid tumors. There were 12 squamous cell carcinomas (SCCs), 11 mast cell tumors (MCTs), 6 hemangiosarcomas (HSAs), 4 adenocarcinomas (ACAs), 3 peripheral nerve sheath tumors (PNSTs), 3 lymphomas, 3 apocrine hidrocystomas (AHCs), and 2 hemangiomas. Cats with MCTs were significantly younger than cats with all other tumor types combined. In contrast, cats with SCCs were significantly older than cats with other tumor types. The HSAs and SCCs were significantly more likely than other tumors to occur in nonpigmented areas. The MCTs, HSAs, AHCs, and hemangiomas did not recur after surgical excision. In contrast, the lymphomas, ACAs, SCCs, and PNSTs frequently recurred and/or resulted in death or euthanasia of the cat. The SCCs were significantly more likely to recur than the MCTs. The average survival time for cats with SCCs was 7.4 months. Although eyelid MCTs have been reported in cats, the prevalence in this study is much higher than previously described.

Squamous cell carcinoma (SCC) is the most common malignant cutaneous and oral neoplasm of cats. Papillomavirus (PV) DNA has been identified in a proportion of feline Bowenoid in situ carcinomas (BISCs), cutaneous SCCs and a single oral SCC, but its exact role in the pathogenesis remains unknown. In humans, it has been suggested that ultraviolet (UV) light and human PV (HPV) may act as cofactors in cutaneous SCC carcinogenesis. Little is known about the influence of UV light on PV prevalence in feline cutaneous lesions, including actinic keratosis (AK). Additionally, PV prevalence in noncutaneous feline lesions, including oral SCC, is largely not known. This study aimed to determine the presence of PV in 84 cats with premalignant and invasive SCC from cutaneous and noncutaneous sites using polymerase chain reaction and to investigate an association with UV light. Papillomaviral DNA was amplified from two of 12 cases of AK, seven of 22 BISCs, nine of 39 cutaneous SCCs and two of 35 non-cutaneous SCCs. Of the PV DNA sequenced, 50% was most similar to HPV of the genus Betapapillomavirus, while the other 50% was most similar to Felis domesticus PV type 2. Exposure to UV was not associated with an increase in PV for cutaneous SCC. The results of this study suggest that in the cat, HPV DNA may be detectible within a higher percentage of squamous lesions than previously demonstrated, UV exposure may not be a confounder for PV presence, and noncutaneous lesions may have a low prevalence of PV.

The related zinc finger transcription factors Slug and Snail modulate epithelial mesenchymal transformation (EMT), the conversion of sessile epithelial cells into migratory fibroblast-like cells. EMT occurs during development, wound healing, and tumor progression. Growth factors, acting through mitogen-activated protein kinase (MAPK) cascades, regulate expression of Slug and Snail. Expression of Snail family transcription factors appears to be elevated in UVR-induced murine squamous cell carcinomas (SCC). We report here that ultraviolet radiation (UVR), which activates MAPK cascades, also stimulates Snail and Slug expression in epidermal keratinocytes. UVR exposure transiently elevated Slug and Snail mRNA expression in human keratinocytes in vitro and mouse epidermis in vivo. This induction was mediated, at least in part, through the ERK and p38 MAPK cascades, as pharmacological inhibition of these cascades partially or completely blocked Slug and Snail induction by UVR. On the other hand, UVR induction of Slug and Snail was enhanced by inhibition of JNK. Slug appears to play a functional role in the acute response of keratinocytes to UVR, as UVR induction of keratin 6 in the epidermis of Slug knockout mice was markedly delayed compared to wild-type mice. Slug and Snail are known to regulate molecules important in the cytoskeleton, intercellular adhesion, cell motility, and apoptosis, thus it seems probable that transiently or persistently elevated expression of these factors fosters the progression of UVR-induced SCC.