The novel lipopolysaccharide-binding protein CRISPLD2 is a critical serum protein to regulate endotoxin function.

LPS is an immunostimulatory component of Gram-negative bacteria. Acting on the immune system in a systemic fashion, LPS exposes the body to the hazard of septic shock. In this study we report that cysteine-rich secretory protein LCCL domain containing 2 (CRISPLD2/Crispld2; human and mouse/rat versions, respectively), expressed by multitissues and leukocytes, is a novel LPS-binding protein. As a serum protein, median CRISPLD2 concentrations in health volunteers and umbilical cord blood samples are 607 microg/ml and 290 microg/ml, respectively. Human peripheral blood granulocytes and mononuclear cells including monocytes, NK cells, and T cells spontaneously release CRISPLD2 (range, 0.2-0.9 microg/ml) and enhance CRISPLD2 secretion (range, 1.5-4.2 microg/ml) in response to stimulation of both LPS and humanized anti-human TLR4-IgA Ab in vitro. CRISPLD2 exhibits significant LPS binding affinity similar to that of soluble CD14, prevents LPS binding to target cells, reduces LPS-induced TNF-alpha and IL-6 production, and protects mice against endotoxin shock. In in vivo experiments, serum Crispld2 concentrations increased in response to a nontoxic dose of LPS and correlated negatively with LPS lethality, suggesting that CRISPLD2 serum concentrations not only are indicators of the degree of a body's exposure to LPS but also reflect an individual's LPS sensitivity.

hBolA, novel non-classical secreted proteins, belonging to different BolA family with functional divergence.

This study reported that all three human BolA proteins (hBolA1, hBolA2, and hBolA3) are novel non-classical secreted proteins identified with bioinformatics and molecular biology experiments. The three BolA fusion proteins with c-Myc tag could be secreted into the culture medium of the transfected Cos-7 cells, although they could not be colocalized with Golgi apparatus. And the secretion of three BolA proteins could not be inhibited after BFA treatment. Furthermore, the secretion was not dependent on its predicted signal peptide. All the experiment results suggested that the secretion was a non-classical export. Phylogenetic analysis showed that the human BolAs belong to three different groups with functional divergence of BolA subfamily, where the different helix-turn-helix motif among hBolA1, hBolA2, and hBolA3 could be responsible for their functional divergence. Our data provided a basis for functional studies of BolA protein family.

hZG16, a novel human secreted protein expressed in liver, was down-regulated in hepatocellular carcinoma.

Here we reported a novel human secreted protein named as hZG16, with a Jacalin domain. Evolution analysis through comparing with the orthologs of other organisms suggested that ZG16 is a conserved gene under the purifying selection (d(N)/d(s)<1) in the evolution. Interestingly, Northern and dot blot analyses showed that hZG16 were highly expressed in adult liver, not in fetal liver, and moderately in gut, including jejunum, ileum, and colon, in which the tissue expression pattern of hZG16 was significantly dissimilar to that of mouse and rat orthologs that were uniquely expressed in spleen and pancreas, respectively. Unexpectedly, hZG16 was markedly down-regulated in hepatocellular carcinoma (HCC) as indicated by RT-PCR, Northern blot analysis and immunohistochemistry staining. However, the tunicamicin treatment and pulse-chase experiments showed that hZG16 protein had a similar molecular function with rZG16 that take part in glycoproteins' secretion in a bus mode.

ZBTB34, a novel human BTB/POZ zinc finger protein, is a potential transcriptional repressor.

BTB/POZ protein family plays a key role in many biological processes both in Drosophila and vertebrates through regulating the transcriptional activities of some downstream genes. Here, we obtained a novel member of human BTB/POZ protein family, named as ZBTB34 (Zinc finger and BTB domain containing 34), which encodes 504 amino acid residues with a BTB/POZ domain at its N-terminus that is similar to the same domain of other known transcription regulators. RT-PCR analysis indicated that ZBTB34 was expressed ubiquitously in most adult human tissues, and whilst immunofluorescence assays showed that ZBTB34 was mainly localized to nucleus. Interestingly, the reporter assay in mammalian cells suggested that ZBTB34 might function as a transcriptional repressor. This present work as the first report about the functional exploration of the novel ZBTB34 gene would be contributed to profound understanding of the transcriptional regulation via BTB/POZ protein family.

[Molecular cloning and characterization of hMGRAP, a human secreted protein with multiple glutamine repeat].

To search for human novel secreted proteins and study their biological functions, using bioinformatical tools and experimental approaches, a novel secreted protein, human hMGRAP (Human Multiple Glutamine Repeat Acidic Protein) was obtained. hMGRAP consists of six coding exons spanning 1547bp of genomic DNA on the human chromosome 7q22.1, which encodes a protein with 248 amino acids. hMGRAP is rich of glutamic acid repeated sequence and the PI is 4.6. The coding sequence of hMGRAP was cloned by PCR method from the cDNA pool composed of nine human tissues. Western blot showed that hMGRAP protein was massively secreted out from the transiently transfected Cos-7 cells. RT-PCR result indicated hMGRAP mRNA was abundantly expressed in testis. In summary, a novel human gene encoding a secreted protein hMGRAP has been screened and cloned, and its biological function may specifically relate to its repeated glutamic acid sequence.