Characterization of Nuclear Localization and SUMOylation of the ATBF1 Transcription Factor in Epithelial Cells
ATBF1/ZFHX3 is a large transcription factor that functions in development, tumorigenesis and other biological processes. ATBF1 is normally localized in the nucleus, but is often mislocalized in the cytoplasm in cancer cells. The mechanism underlying the mislocalization of ATBF1 is unknown. In this study, we analyzed the nuclear localization of ATBF1, and found that ectopically expressed ATBF1 formed nuclear body (NB)-like dots in the nucleus, some of which indeed physically associated with promyelocytic leukemia (PML) NBs. We also defined a 3-amino acid motif, KRK2615-2617, as the nuclear localization signal (NLS) for ATBF1. Interestingly, diffusely distributed nuclear SUMO1 proteins were sequestered into ATBF1 dots, which could be related to ATBF1's physical association with PML NBs, known SUMOylation hotspots. Furthermore, ATBF1 itself was SUMOylated. ATBF1 SUMOylation occurred at more than 3 lysine residues including K2349, K2806 and K3258 and was nuclear specific. Finally, the PIAS3 SUMO1 E3 ligase, which interacts with ATBF1 directly, diminished rather than enhanced ATBF1 SUMOylation, preventing the co-localization of ATBF1 with SUMO1 in the nucleus. These findings suggest that nuclear localization and SUMOylation are important for the transcription factor function of ATBF1, and that ATBF1 could cooperate with PML NBs to regulate protein SUMOylation in different biological processes.

title: Heterozygous deletion of Atbf1 by the Cre-loxP system in mice causes preweaning mortality.
abstract: ATBF1 is a large nuclear protein that contains multiple zinc-finger motifs and four homeodomains. In mammals, ATBF1 regulates differentiation, and its mutation and/or downregulation is involved in tumorigenesis in several organs. To gain more insight into the physiological functions of ATBF1, we generated and validated a conditional allele of mouse Atbf1 in which exons 7 and 8 were flanked by loxP sites (Atbf1(flox) ). Germline deletion of a single Atbf1 allele was achieved by breeding to EIIa-cre transgenic mice, and Atbf1 heterozygous mice displayed reduced body weight, preweaning mortality, increased cell proliferation, and attenuated cytokeratin 18 expression, indicating haploinsufficiency of Atbf1. Floxed Atbf1 mice will help us understand such biological processes as neuronal differentiation and tumorigenesis.

title: Influence of N-substituent and solvent on internal conversion in 1-aminonaphthalenes.
abstract: N-Methyl-N-methoxycarbonylmethyl-1-aminonaphthalene (MMAN) and N,N-dimethoxycarbonylmethyl-1-aminonaphthalene (DCAN) were synthesized and their fluorescence and absorption spectra in solvents of varied polarity were investigated and compared to those of 1-dimethylaminonaphthylene (DMAN). The data were discussed in terms of the N-substitution effects on the internal conversion (IC). Results showed that the IC was enhanced when the N-methyl group in DMAN was replaced by N-CH2CO2CH3.

title: A novel biosensor for bovine serum albumin based on fluorescent self-assembled sandwich bilayers.
abstract: Fluorescent dyes butyl rhodamine B were assembled via a DL-cystenine intermediate onto quartz wafers whose surface had first adsorbed gold nanoparticles. Hence self-assembled sandwich bilayers with nanocomposite structure were constructed which can be used as a biosensor for bovine serum albumin. The biosensor-based self-assembled monolayers (SAMs) are regenerable and have high sensitivity, five orders of magnitude higher than that of bulk solution phase sensing. The effects of existing forms of dyes on the fluorescence spectra of bilayers in the presence of bovine serum albumin were investigated.

title: In pursuit of leucine-rich repeat-containing G protein-coupled receptor-5 regulation and function in the uterus.
abstract: Leucine-rich repeat-containing G protein-coupled receptor (LGR)-5 is a recently identified marker of stem cells in adult intestinal epithelium and hair follicles. Because of this characteristic, we studied the status of Lgr5 expression in the mouse uterus under various conditions. Lgr5 is highly expressed in the uterine epithelium of immature mice and is dramatically down-regulated after the mice resume estrous cycles. Surprisingly, whereas its expression is up-regulated in uteri of ovariectomized mice, the expression is down-regulated by estrogen and progesterone via their cognate nuclear receptors, estrogen receptor-alpha and progesterone receptor, respectively. Using a mouse endometrial cancer model, we also found that Lgr5 is highly expressed in the epithelium during the initial stages of tumorigenesis but is remarkably down-regulated in fully developed tumors. Lgr5 is a downstream target of Wnt signaling in the intestine. Genetic evidence shows that either excessive or absence of Wnt signaling dampens Lgr5 expression in the uterus. Collectively, our results show that Lgr5 expression in the mouse uterine epithelium is unique and dynamically regulated under various physiological and pathological states of the uterus, suggesting that this orphan receptor has important functions in uterine biology. However, identifying definitive uterine function of LGR5 will require further investigation using conditional deletion of uterine Lgr5 because systemic deletion of this gene is neonatally lethal.

title: Performance optimization in electric field gradient focusing.
abstract: Electric field gradient focusing (EFGF) is a technique used to simultaneously separate and concentrate biomacromolecules, such as proteins, based on the opposing forces of an electric field gradient and a hydrodynamic flow. Recently, we reported EFGF devices fabricated completely from copolymers functionalized with poly(ethylene glycol), which display excellent resistance to protein adsorption. However, the previous devices did not provide the predicted linear electric field gradient and stable current. To improve performance, Tris-HCl buffer that was previously doped in the hydrogel was replaced with a phosphate buffer containing a salt (i.e., potassium chloride, KCl) with high mobility ions. The new devices exhibited stable current, good reproducibility, and a linear electric field distribution in agreement with the shaped gradient region design due to improved ion transport in the hydrogel. The field gradient was calculated based on theory to be approximately 5.76 V/cm(2) for R-phycoerythrin when the applied voltage was 500 V. The effect of EFGF separation channel dimensions was also investigated; a narrower focused band was achieved in a smaller diameter channel. The relationship between the bandwidth and channel diameter is consistent with theory. Three model proteins were resolved in an EFGF channel of this design. The improved device demonstrated 14,000-fold concentration of a protein sample (from 2 ng/mL to 27 microg/mL).

title: Kruppel-like factor 5 (KLF5) is critical for conferring uterine receptivity to implantation
abstract: A blastocyst will implant only when the uterus becomes receptive. Following attachment, luminal epithelial cells undergo degeneration at the site of the blastocyst. Although many genes critical for uterine receptivity are primarily regulated by ovarian hormones, Kruppel-like factor 5 (KLF5), a zinc finger-containing transcription factor, is persistently expressed in epithelial cells independently of ovarian hormones. Loss of uterine Klf5 causes female infertility due to defective implantation. Cox2 is normally expressed in the luminal epithelium and stroma at the site of blastocyst attachment, but luminal epithelial COX2 expression is absent with loss of Klf5. This is associated with the retention of the epithelium around the implantation chamber with arrested embryonic growth. These results suggest that Klf5 is indispensable for normal implantation.