scholarly journals Immunocytochemical Localization of Olfactory-signaling Molecules in Human and Rat Spermatozoa

2020 ◽  
Vol 68 (7) ◽  
pp. 491-513 ◽  
Author(s):  
Yuliya Makeyeva ◽  
Christopher Nicol ◽  
William L. Ledger ◽  
David K. Ryugo
Keyword(s):  
Sensory Neurons ◽  
Expression Patterns ◽  
Signal Transduction Pathway ◽  
Olfactory Receptors ◽  
Signaling Cascades ◽  
Transduction Pathway ◽  
Olfactory Marker Protein ◽  
Functional Maturation ◽  
Motile Cells ◽  
Second Messenger Signaling

Expression of olfactory receptors (ORs) in non-olfactory tissues has been widely reported over the last 20 years. Olfactory marker protein (OMP) is highly expressed in mature olfactory sensory neurons (mOSNs) of the olfactory epithelium. It is involved in the olfactory signal transduction pathway, which is mediated by well-conserved components, including ORs, olfactory G protein (Golf), and adenylyl cyclase 3 (AC3). OMP is widely expressed in non-olfactory tissues with an apparent preference for motile cells. We hypothesized that OMP is expressed in compartment-specific locations and co-localize with an OR, Golf, and AC3 in rat epididymal and human-ejaculated spermatozoa. We used immunocytochemistry to examine the expression patterns of OMP and OR6B2 (human OR, served as positive olfactory control) in experimentally induced modes of activation and determine whether there are any observable differences in proteins expression during the post-ejaculatory stages of spermatozoal functional maturation. We found that OMP was expressed in compartment-specific locations in human and rat spermatozoa. OMP was co-expressed with Golf and AC3 in rat spermatozoa and with OR6B2 in all three modes of activation (control, activated, and hyperactivated), and the mode of activation changed the co-expression pattern in acrosomal-reacted human spermatozoa. These observations suggest that OMP expression is a reliable indicator of OR-mediated chemoreception, may be used to identify ectopically expressed ORs, and could participate in second messenger signaling cascades that mediate fertility.

Differential Expression of Genes Associated with Toll-Like Receptor-4 (TLR4) Signal Transduction Pathway in Lipopolysaccharide (LPS)-Activated Cord Blood (CB) vs. Adult Peripheral Blood (APB) Monocyte (Mo)-Derived Dendritic Cells (DC).

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 3444-3444
Author(s):  
Hong Jiang ◽  
Mary Brigid Bradley ◽  
Carmella van de Ven ◽  
Prakash Satwani ◽  
Laxmi Baxi ◽  
...  
Keyword(s):  
Gene Expression ◽  
Signal Transduction ◽  
Protein Expression ◽  
Expression Patterns ◽  
Signal Transduction Pathway ◽  
Gene Expression Patterns ◽  
Transduction Pathway ◽  
Rt Pcr ◽  
Cell Surface Molecule ◽  
Expression Of Genes

Abstract LPS activates immature DC via TLR4 and induces maturation of DC for initiating antigen presenting activity (Medzhitov; Nat Rev Immunol 2001). We have previously demonstrated decreased gene expression and protein production of IL-12, IL-15, IL-18 in activated CB MNC and decreased DC MLR (Lee/Cairo, Blood 1996; Qian/Cairo, Blood 1997; Wu/Cairo, Blood 100:3668 p51b 2002). Recently, we have identified differential gene expression patterns including differential immunoregulatory and chemokine genes in LPS-CB vs APB Mo by microarray (Jiang/Cairo, J. Immunol 2004). Since the myeloid lineage DC is derived from Mo, we sought to determine in LPS activated CB vs. APB DC, differential expressed genes that associate with TLR4-mediated signaling pathway. Briefly, Mo were purified from fresh CB or APB and cultured for 7 days with GM-CSF & IL-4 [immature DC (iDC)] and LPS [mature DC (mDC)]. Aliquots from iDC and mDC were analyzed for DC immunophenotype, morphology and DC allogeneic antigen activity. mRNA was isolated, reverse transcripted to cDNA, labeled & hybridized to oligonucleotides (Affymetrix, U133A). Data was analyzed by MAS 5.0 (Affymetrix) and GeneSpring 5.0 software (Silicon Genetics). Several genes were analyzed by RT-PCR (One-Step SuperScript, Invitrogen) and protein expression was analyzed by Western Blot (Bio-Rad). Inverted microscopy demonstrated DC mature morphology at day 8 and flow cytometry demonstrated decreased CD14 and increased CD83 expression in CB & APB mDC. We also demonstrated significant increase in the allogeneic stimulatory effects on CD4+ T cells in APB vs. CB mDC. The microarray analysis demonstrated a significant decreased gene expression of TLR4 [3 fold (F)] and CD14 (2.1 F) (p<0.05) in CB vs APB-DC. We further identified LPS significantly induced increased expression of TLR4 downstream signaling molecular genes such as MAPKKK, NF-kB and TANK in APB compared to CB mDC (3–8 F) (p<0.05). There were also significant amplifications of a variety of other gene categories in LPS activated APB vs CB mDC (p<0.05) including cell surface molecule CD80 (3.7F) and IL-2Ra (5.3 F), cytokine IL-23 (3.5F) & IL-12 (13 F), signal transduction STAT1 (3.4F) & IRF-7 (7.7 F), and immunoregulatory TNFSF10 (12F) & ISG20 (39F). Gene expression of NF-kB1, TRAF1 & IRF-7 by RT-PCR demonstrated an increased expression in LPS-APB vs CB mDC and were compatible with microarray. Moreover, Western analysis of IRF-7 demonstrated increased protein expression in LPS-APB vs CB mDC. In summary, we have identified decreased gene expression patterns in LPS-CB vs APB DC, especially those in the TLR4 signal transduction pathway (MAP3K, TRAF, TANK & NF-kB), and suggest these differentially expressed genes may enhance the activation of TLR4 pathway in LPS-APB vs CB DC, resulting in differential regulation of CB vs APB DC antigen presentation capacities. Furthermore, these decreased expressed genes in other molecular categories (e.g.IL-23, IFNg, IL6, CD80, STAT1, IRF-7, SOCS3) in LPS-CB vs APB DC may be partially responsible for differential innate and adaptive immune function of CB vs APB. Moreover, the differential regulated expression of genes may in part help to explain reduced incidence of severe aGVHD, delay in immune reconstitution and/or increased infectious mortality following HLA disparate UCBT.

scholarly journals SdeK, a Histidine Kinase Required for Myxococcus xanthus Development

2001 ◽  
Vol 183 (12) ◽  
pp. 3589-3596 ◽  
Author(s):  
Jeffrey S. Pollack ◽  
Mitchell Singer
Keyword(s):  
Signal Transduction ◽  
Histidine Kinase ◽  
Developmental Process ◽  
Expression Patterns ◽  
Myxococcus Xanthus ◽  
Signal Transduction Pathway ◽  
Transduction Pathway ◽  
Protein Activity

ABSTRACT The sdeK gene is essential to the Myxococcus xanthus developmental process. We reported previously, based on sequence analysis (A. G. Garza, J. S. Pollack, B. Z. Harris, A. Lee, I. M. Keseler, E. F. Licking, and M. Singer, J. Bacteriol. 180:4628–4637, 1998), that SdeK appears to be a histidine kinase. In the present study, we have conducted both biochemical and genetic analyses to test the hypothesis that SdeK is a histidine kinase. An SdeK fusion protein containing an N-terminal polyhistidine tag (His-SdeK) displays the biochemical characteristics of a histidine kinase. Furthermore, histidine 286 of SdeK, the putative site of phosphorylation, is required for both in vitro and in vivo protein activity. The results of these assays have led us to conclude that SdeK is indeed a histidine kinase. The developmental phenotype of a ΔsdeK1 strain could not be rescued by codevelopment with wild-type cells, indicating that the defect is not due to the mutant's inability to produce an extracellular signal. Furthermore, the ΔsdeK1 mutant was found to produce both A- and C-signal, based on A-factor and codevelopment assays with acsgA mutant, respectively. The expression patterns of several Tn5lacZ transcriptional fusions were examined in the ΔsdeK1-null background, and we found that all C-signal-dependent fusions assayed also required SdeK for full expression. Our results indicate that SdeK is a histidine kinase that is part of a signal transduction pathway which, in concert with the C-signal transduction pathway, controls the activation of developmental-gene expression required to progress past the aggregation stage.

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