The United Kingdom Primary Immune Deficiency (UKPID) registry 2012 to 2017.

This is the second report of the United Kingdom Primary Immunodeficiency (UKPID) registry. The registry will be a decade old in 2018 and, as of August 2017, had recruited 4758 patients encompassing 97% of immunology centres within the United Kingdom. This represents a doubling of recruitment into the registry since we reported on 2229 patients included in our first report of 2013. Minimum PID prevalence in the United Kingdom is currently 5·90/100 000 and an average incidence of PID between 1980 and 2000 of 7·6 cases per 100 000 UK live births. Data are presented on the frequency of diseases recorded, disease prevalence, diagnostic delay and treatment modality, including haematopoietic stem cell transplantation (HSCT) and gene therapy. The registry provides valuable information to clinicians, researchers, service commissioners and industry alike on PID within the United Kingdom, which may not otherwise be available without the existence of a well-established registry.

An activation switch in the ligand binding pocket of the C5a receptor.

Although agonists are thought to occupy binding pockets within the seven-helix core of serpentine receptors, the topography of these binding pockets and the conformational changes responsible for receptor activation are poorly understood. To identify the ligand binding pocket in the receptor for complement factor 5a (C5aR), we assessed binding affinities of hexapeptide ligands, each mutated at a single position, for seven mutant C5aRs, each mutated at a single position in the putative ligand binding site. In ChaW (an antagonist) and W5Cha (an agonist), the side chains at position 5 are tryptophan and cyclohexylalanine, respectively. Comparisons of binding affinities indicated that the hexapeptide residue at this position interacts with two C5aR residues, Ile-116 (helix III) and Val-286 (helix VII); in a C5aR model these two side chains point toward one another. Both the I116A and the V286A mutations markedly increased binding affinity of W5Cha but not that of ChaW. Moreover, ChaW, the antagonist hexapeptide, acted as a full agonist on the I116A mutant. These results argue that C5aR residues Ile-116 and Val-286 interact with the side chain at position 5 of the hexapeptide ligand to form an activation switch. Based on this and previous work, we present a docking model for the hexapeptide within the C5aR binding pocket. We propose that agonists induce a small change in the relative orientations of helices III and VII and that these helices work together to allow movement of helix VI away from the receptor core, thereby triggering G protein activation.

Leukocytes navigate by compass: roles of PI3Kgamma and its lipid products.

Morphologic polarity is necessary for the motility of mammalian cells. In leukocytes responding to a chemoattractant, this polarity is regulated by activities of small Rho guanosine triphosphatases (Rho GTPases) and the phosphoinositide 3-kinases (PI3Ks). Moreover, in neutrophils, lipid products of PI3Ks appear to regulate activation of Rho GTPases, are required for cell motility and accumulate asymmetrically to the plasma membrane at the leading edge of polarized cells. By spatially regulating Rho GTPases and organizing the leading edge of the cell, PI3Ks and their lipid products could play pivotal roles not only in establishing leukocyte polarity but also as compass molecules that tell the cell where to crawl.

Polarization of chemoattractant receptor signaling during neutrophil chemotaxis.

Morphologic polarity is necessary for chemotaxis of mammalian cells. As a probe of intracellular signals responsible for this asymmetry, the pleckstrin homology domain of the AKT protein kinase (or protein kinase B), tagged with the green fluorescent protein (PHAKT-GFP), was expressed in neutrophils. Upon exposure of cells to chemoattractant, PHAKT-GFP is recruited selectively to membrane at the cell's leading edge, indicating an internal signaling gradient that is much steeper than that of the chemoattractant. Translocation of PHAKT-GFP is inhibited by toxin-B from Clostridium difficile, indicating that it requires activity of one or more Rho guanosine triphosphatases.

Dynamics of a chemoattractant receptor in living neutrophils during chemotaxis.

Persistent directional movement of neutrophils in shallow chemotactic gradients raises the possibility that cells can increase their sensitivity to the chemotactic signal at the front, relative to the back. Redistribution of chemoattractant receptors to the anterior pole of a polarized neutrophil could impose asymmetric sensitivity by increasing the relative strength of detected signals at the cell's leading edge. Previous experiments have produced contradictory observations with respect to receptor location in moving neutrophils. To visualize a chemoattractant receptor directly during chemotaxis, we expressed a green fluorescent protein (GFP)-tagged receptor for a complement component, C5a, in a leukemia cell line, PLB-985. Differentiated PLB-985 cells, like neutrophils, adhere, spread, and polarize in response to a uniform concentration of chemoattractant, and orient and crawl toward a micropipette containing chemoattractant. Recorded in living cells, fluorescence of the tagged receptor, C5aR-GFP, shows no apparent increase anywhere on the plasma membrane of polarized and moving cells, even at the leading edge. During chemotaxis, however, some cells do exhibit increased amounts of highly folded plasma membrane at the leading edge, as detected by a fluorescent probe for membrane lipids; this is accompanied by an apparent increase of C5aR-GFP fluorescence, which is directly proportional to the accumulation of plasma membrane. Thus neutrophils do not actively concentrate chemoattractant receptors at the leading edge during chemotaxis, although asymmetrical distribution of membrane may enrich receptor number, relative to adjacent cytoplasmic volume, at the anterior pole of some polarized cells. This enrichment could help to maintain persistent migration in a shallow gradient of chemoattractant.

Galphai is not required for chemotaxis mediated by Gi-coupled receptors.

Pertussis toxin inhibits chemotaxis of neutrophils by preventing chemoattractant receptors from activating trimeric G proteins in the Gi subfamily. In HEK293 cells expressing recombinant receptors, directional migration toward appropriate agonist ligands requires release of free G protein betagamma subunits and can be triggered by agonists for receptors coupled to Gi but not by agonists for receptors coupled to two other G proteins, Gs and Gq. Because activation of any G protein presumably releases free Gbetagamma, we tested the hypothesis that chemotaxis also requires activated alpha subunits (Galphai) of Gi proteins. HEK293 cells were stably cotransfected with the Gi-coupled receptor for interleukin-8, CXCR1, and with a chimeric Galpha, Galphaqz5, which resembles Galphai in susceptibility to activation by Gi-coupled receptors but cannot regulate the Galphai effector, adenylyl cyclase. These cells, unlike cells expressing CXCR1 alone, migrated toward interleukin-8 even after treatment with pertussis toxin, which prevents activation of endogenous Galphai but not that of Galphaqz5. We infer that chemotaxis does not require activation of Galphai. Because chemotaxis is mediated by Gbetagamma subunits released when Gi-coupled receptors activate Galphaqz5, but not when Gq- or Gs-coupled receptors activate their respective G proteins, we propose that Gi-coupled receptors transmit a necessary chemotactic signal that is independent of Galphai.

A Gsalpha mutant designed to inhibit receptor signaling through Gs.

Hormonal signals activate trimeric G proteins by substituting GTP for GDP bound to the G protein alpha subunit (Galpha), thereby generating two potential signaling molecules, Galpha-GTP and free Gbetagamma. The usefulness of dominant negative mutations for investigating Ras and other monomeric G proteins inspired us to create a functionally analogous dominant negative Galpha mutation. Here we describe a mutant alpha subunit designed to inhibit receptor-mediated hormonal activation of Gs, the stimulatory regulator of adenylyl cyclase. To construct this mutant, we introduced into the alpha subunit (alphas) of Gs three separate mutations chosen because they impair alphas function in complementary ways: the A366S mutant reduces affinity of alphas for binding GDP, whereas the G226A and E268A mutations impair the protein's ability to bind GTP and to assume an active conformation. The triple mutant robustly inhibits (by up to 80%) Gs-dependent hormonal stimulation of adenylyl cyclase in cultured cells. Inhibition is selective in that it does not affect cellular responses to expression of a constitutively active alphas mutant (alphas-R201C) or to agonists for receptors that activate Gq or Gi. This alphas triple mutant and cognate Galpha mutants should provide specific tools for dissection of G protein-mediated signals in cultured cells and transgenic animals.

Second site suppressor mutations of a GTPase-deficient G-protein alpha-subunit. Selective inhibition of Gbeta gamma-mediated signaling.

G proteins transmit signals from cell surface receptors to intracellular effectors. The intensity of the signal is governed by the rates of GTP binding (leading to subunit dissociation) and hydrolysis. Mutants that cannot hydrolyze GTP (e.g. GsalphaQ227L, Gi2alphaQ205L) are constitutively activated and can lead to cell transformation and cancer. Here we have used a genetic screen to identify intragenic suppressors of a GTPase-deficient form of the Galpha in yeast, Gpa1(Q323L). Sequencing revealed second-site mutations in three conserved residues, K54E, R327S, and L353Delta (codon deletion). Each mutation alone results in a complete loss of the beta gamma-mediated mating response in yeast, indicating a dominant-negative mode of inhibition. Likewise, the corresponding mutations in a mammalian Gi2alpha (K46E, R209S, L235Delta) lead to inhibition of Gbeta gamma-mediated mitogen-activated protein (MAP) kinase phosphorylation in cultured cells. The most potent of these beta gamma inhibitors (R209S) has no effect on Gi2alpha-mediated regulation of adenylyl cyclase. Despite its impaired ability to release beta gamma, purified recombinant Gpa1(R327S) is fully competent to bind and hydrolyze GTP. These mutants will be useful for uncoupling Gbeta gamma- and Galpha-mediated signaling events in whole cells and animals. In addition, they serve as a model for drugs that could directly inhibit G protein activity and cell transformation.

Conditional activation defect of a human Gsalpha mutant.

Hormonal signals activate trimeric G proteins by promoting exchange of GTP for GDP bound to the G protein's alpha subunit (Galpha). Here we describe a point mutation that impairs this activation mechanism in the alpha subunit of Gs, producing an inherited disorder of hormone responsiveness. Biochemical analysis reveals an activation defect that is paradoxically intensified by hormonal and other stimuli. By substituting histidine for a conserved arginine residue, the mutation removes an internal salt bridge (to a conserved glutamate) that normally acts as an intramolecular hasp to maintain tight binding of the gamma-phosphate of GTP. In its basal, unperturbed state, the mutant alphas binds guanosine 5'-[gamma-thio]triphosphate (GTP[gammaS]), a GTP analog, slightly less tightly than does normal alphas, but (in the GTP[gammaS]-bound form) can stimulate adenylyl cyclase. The activation defect becomes prominent only under conditions that destabilize binding of guanine nucleotide (receptor stimulation) or impair the ability of alphas to bind the gamma-phosphate of GTP (cholera toxin, AlF4- ion). Although GDP release is usually the rate-limiting step in nucleotide exchange, the biochemical phenotype of this mutant alphas indicates that efficient G protein activation by receptors and other stimuli depends on the ability of Galpha to clasp tightly the GTP molecule that enters the binding site.

Testicular and epididymal sperm in a microinjection program: methods of retrieval and results.

OBJECTIVE: To describe the methods of collection and laboratory preparation of epididymal and testicular sperm; to compare the fertilization and pregnancy rates; and to establish prognostic factors. DESIGN: Retrospective analysis. SETTING: Academic reproductive medicine clinic. PATIENT(S): One hundred twelve consecutive microinjection cycles in 80 patients using either epididymal or testicular sperm. INTERVENTION(S): Sperm were collected by microepididymal sperm aspiration, open testicular biopsy, or fine needle tissue aspiration testicular biopsy. MAIN OUTCOME MEASURE(S): Fertilization rate, implantation, and clinical pregnancy rates. RESULT(S): The fertilization rate was higher with epididymal sperm (67%) than with testicular sperm (50%). Implantation rates (fetal hearts per embryo, testicular: 11%, epididymal: 8%) and pregnancy rates (clinical pregnancy per oocyte collection procedure, testicular: 25%, epididymal: 29%) were not significantly different with epididymal and testicular sperm. Multiple regression analysis showed that normal fertilization rates were significantly lower with testicular sperm, immotile sperm, and severe spermatogenic disorders. CONCLUSION(S): Although fertilization rates are significantly lower with testicular sperm, higher implantation rates resulted in equivalent pregnancy rates. Thus, testicular aspiration of sperm for intracytoplasmic sperm injection is a simple, inexpensive method of sperm retrieval in cases of azoospermia resulting from genital tract obstruction or severe spermatogenic disorder.

Testicular aspiration of sperm for intracytoplasmic sperm injection: a novel treatment for ejaculatory failure on the day of oocyte retrieval.

OBJECTIVE: To present a successful outcome of an IVF cycle complicated by failure to produce a sperm sample on the morning of oocyte retrieval. DESIGN: Case report. SETTING: Hospital-based IVF clinic. PATIENT: A couple suffering secondary infertility undergoing their first IVF and intracytoplasmic sperm injection (ICSI) cycle. INTERVENTION: Fine-needle testicular biopsy for sperm retrieval. MAIN OUTCOME MEASURES: Sperm retrieval, fertilization, pregnancy. RESULTS: Sufficient spermatozoa were obtained for ICSI. Ten metaphase II oocytes were injected with two embryos being transferred fresh and four cryopreserved. A single fetal heart was confirmed at 6-week sonar. CONCLUSION: Testicular aspiration of sperm for ICSI is a simple, inexpensive, and effective means of rescuing an IVF cycle in which the male partner unexpectedly has been unable to produce a sperm sample on the morning of oocyte collection.

Variations in the screening history and appropriateness of management of cervical cancer in South East England.

In seven health districts in southern England, an audit of the management of cervical cancer compared with regionally developed guidelines was undertaken between 1988 and 1991. Four hundred and sixty-nine regional residents were treated in the study district hospitals. 73 (15.6%) women were appropriately staged, with increasing likelihood of appropriate staging investigations observed with higher stages (P < 0.0001) and type of hospital [Teaching 23 (21%), Non-Teaching with oncology support 11 (11.5%), Non-teaching 4 (7%), P < 0.0001] but with no change over the study period. There was no significant trend in the proportion of women treated appropriately over time, with 270 (59%) being appropriately treated, 91 (20%) under-treated and 98 (21%) over-treated overall. Appropriateness of treatment increased with higher stages (P < 0.0001) and hospital workload for cancer of the cervix (P = 0.038). Multivariable analysis indicated that survival independently and significantly decreased with age and stage, under-treatment and in cases where lymph nodes were involved or not examined. There was no change in the appropriateness of management over the 4 years, with high levels of inappropriate care. Survival was not only influenced by biological and demographic factors, but by inappropriate care.

Variations in the management and survival of women under 50 years with breast cancer in the South East Thames region.

A retrospective, population-based study was undertaken to determine variations in the management of women aged less than 50 years with primary breast cancer in different hospital settings and the influence of these variations on survival. A total of 1757 women who were resident in the South East Thames Health Region aged less than 50 years at the time of diagnosis of breast cancer and who presented during a 5 year period (January 1984 to December 1988) were recorded by the Thames Cancer Registry. The hospitals at which primary surgery was undertaken were categorised as teaching or non-teaching hospitals. The non-teaching hospitals were grouped according to the mean number of patients treated annually during the study period (< or = 2, 3-9, > or = 10 each year). The following factors were compared between these groups: age, extent of disease, tumour morphology, extent of primary surgery (mastectomy vs less than mastectomy), use of axillary surgery (any vs none) and use of systemic adjuvant therapy. Survival rates for the different groups were compared. Registration rates did not differ significantly between health districts. A total of 1485 (85%) women underwent surgery in over 90 different hospitals. In 1324 (86%) of these cases the surgery was undertaken in a total of 42 NHS hospitals within SE Thames Health Region or in seven teaching hospitals in adjacent regions. Mastectomy rates decreased from 52% in 1984 to 28% in 1988 (P<0.0001), but were consistently higher in teaching hospitals (P=0.01). The use of any form of axillary surgery decreased from 49% to 36% over the 5 year period (P=0.003), with significantly lower rates of axillary surgery being performed in non-teaching hospitals (P<0.0001). The proportion of cases recorded as having non-specific morphology was higher in nonteaching than in teaching hospitals (P<0.0001). On multivariate analysis survival was significantly (P<0.001) influenced by stage and tumour histology. Among patients who underwent surgery, the type of hospital in which this was undertaken did not appear to influence survival significantly. This analysis of routine cancer registry data indicates that patients were widely dispersed in a large number of different hospitals and that there were marked variations in practice according to the type of hospital to which patients presented. The treatments provided were frequently at variance with those recommended at a consensus conference held during the study period, particularly in relation to the use of axillary surgery and adjuvant systemic therapy. The way in which services are currently provided may hamper the delivery of appropriate management and comprehensive support. These data thus have implications for the purchasing and provision of services for this common condition.

Gi2-mediated activation of the MAP kinase cascade.

The Gi class of heterotrimeric G proteins has been implicated in transmitting mitogenic signals from a variety of seven-transmembrane domain receptors. In addition, the alpha subunit of Gi2 (alpha i2) is oncogenic when mutated to a constitutively active form (gip2). The mechanism by which Gi2 stimulates cellular proliferation is unknown, but is believed to involve activation of the mitogen-activated protein kinase (MAPK) signaling cascade. To study Gi2 activation of the cascade, we transiently expressed a mutant, pertussis toxin (PTX)-resistant alpha i2 in Chinese hamster ovary cells. After PTX treatment of these cells, Gi-coupled receptors specifically activated PTX-resistant Gi2 without activating other Gi proteins. Receptor-mediated activation of Gi2 led to activation of MAPK and its upstream activator, MAPK/ERK-activating kinase (MEK). Activation of MAPK and MEK by Gi2 was blocked by expression of a dominant-negative mutant of Ras. Gi2 activation did not, however, detectably increase the proportion of Ras protein in the GTP-bound form. Additional experiments suggest that Gi2 stimulates the MAPK pathway, at least in part, by mechanisms that involve release of its beta gamma subunit, as well as activation of phosphatidylinositol-3 kinase.

Pregnancies after intracytoplasmic injection of sperm collected by fine needle biopsy of the testis.

OBJECTIVE: To investigate the use of intracytoplasmic sperm injection with sperm collected by fine needle biopsy of the testis as a treatment for male genital tract obstruction. DESIGN: Sperm isolated from a fine needle biopsy of the testis were used to inseminate oocytes by intracytoplasmic sperm injection. SETTING: A hospital-based tertiary referral infertility service. PATIENTS: Case studies of two couples in whom the male partner had a genital tract obstruction. MAIN OUTCOME MEASURES: Fertilization and pregnancy. RESULTS: In the first case, 9 oocytes fertilized normally out of 13 injected. After the transfer of fresh (one cycle) and frozen (four cycles) embryos, a single intrauterine fetal heart pregnancy was achieved. In the second case, five oocytes fertilized normally from nine oocytes injected; two embryos were transferred fresh and three were frozen. A single fetal heart intrauterine gestation was obtained after the initial transfer of two fresh embryos. CONCLUSIONS: A high normal fertilization rate and pregnancies are possible with intracytoplasmic sperm injection using sperm collected directly from the testis. Sperm retrieval by fine needle biopsy offers a viable alternative to microsurgical aspiration and is also suitable for treating patients with intratesticular blockage.

Differential effects on cAMP on the MAP kinase cascade: evidence for a cAMP-insensitive step that can bypass Raf-1.

Because cAMP exerts opposite effects on cell proliferation in different cell types, we undertook to study its effect on the mitogen-activated protein kinase (MAPK) pathway in three cell lines (Rat-1, Swiss-3T3, and COS-7) chosen for their different mitogenic responses to cAMP. We measured the effect of cAMP on MAPK, MEK, and Raf-1 activities after stimulation by agonists acting through a tyrosine kinase receptor (epidermal growth factor) or a G protein-coupled receptor (lysophosphatidic acid). In Rat-1 cells we found that cAMP strongly inhibited all three activities (MAPK, MEK, and Raf-1), in good agreement with its effect on cell proliferation in these cells. In Swiss-3T3 and COS-7 cells, on the contrary, cAMP did not inhibit epidermal growth factor- and lysophosphatidic acid-induced stimulation of MAPK and MEK activities, and even stimulated MAPK activity slightly on its own. Again these results are in good agreement with the proliferative effect of cAMP in Swiss-3T3 cells. Raf-1 activity on the hand, was inhibited by cAMP in Swiss-3T3 and COS-7 as it was in Rat-1 cells. This result indicates that signaling pathways in Swiss-3T3 and COS-7 cells can activate MEK and MAPK in a Raf-1-independent and cAMP-insensitive manner. Our results add to growing evidence for the existence of Ras- and/or Raf-1-independent pathways leading to MEK and MAPK activation.

Mutant alpha subunit of Gz transforms Swiss 3T3 cells.

Many hormones and neurotransmitters induce cell proliferation by regulating signaling pathways controlled by heterotrimeric G proteins. Mutations that activate the alpha subunits of Gs and Gi2 produce the gsp and gip2 oncogenes that are found in certain human endocrine tumors. Similar mutations have conferred on other G alpha subunits the ability to promote neoplastic transformation in cultured mammalian cells. Gz, a G protein whose normal signaling function is poorly understood, shares with Gi2 the ability to inhibit adenylyl cyclase. We asked whether mutationally activated alpha z can stimulate cell proliferation in a cell line in which stimulation adenylyl cyclase is mitogenic, Swiss 3T3 cells. Stable expression of alpha z-Q205L in Swiss 3T3 cells induced focus formation, a faster growth rate with a higher saturation density, anchorage-independent growth in soft agar, and increased [3H]thymidine incorporation in the absence of growth factors. alpha z-Q205L produced a similar but less extensively transformed phenotype in NIH3T3 cells--increased saturation density in culture, a smaller number of foci and few colonies in soft agar. Stimulation of thymidine incorporation by alpha z-Q205L in Swiss 3T3 cells was increased by co-treatment with cholera toxin, a stimulator of adenylyl cyclase. Taken together, our results indicate that alpha z stimulates one or more mitogenic pathways in Swiss 3T3 cells, and that effectiveness of these mitogenic pathways does not require reducing the concentration of cellular cAMP.

Fatty acylation of alpha z. Effects of palmitoylation and myristoylation on alpha z signaling.

As the first step in an investigation of roles played by fatty acylation of G protein alpha chains in membrane targeting and signal transmission, we inserted monoclonal antibody epitopes, hemagglutinin (HA) or Glu-Glu (EE), at two internal sites in three alpha subunits. At site I, only HA-tagged alpha q and alpha z functioned normally. alpha s, alpha q, and alpha z subunits tagged at site II with the EE epitope showed normal expression, membrane localization, and signaling activity. Using epitope-tagged alpha z, we investigated effects of mutations in sites for fatty acylation. Mutational substitution of Ala for Gly2 (G2A) prevented incorporation of myristate and decreased but did not abolish incorporation of palmitate. Substitution of Ala for Cys3 (C3A) prevented incorporation of palmitate but had no effect on incorporation of myristate. Substitution of Ala for both Gly2 and Cys3 (G2AC3A) prevented incorporation of both myristate and palmitate. All three mutations substantially disrupted association of alpha z with the particulate fraction. Gz-mediated inhibition of adenylyl cyclase, triggered by activation of the D2-dopamine receptor, was, respectively, abolished (G2AC3A), impaired (G2A), and enhanced (C3A). Constitutive inhibition of adenylyl cyclase by alpha z was unchanged (G2AC3A), strongly diminished (G2A), or strongly enhanced (C3A). A nonacylated, mutationally activated alpha z mutant inhibited adenylyl cyclase, although less potently than normally acylated, mutationally activated alpha z. From these findings we conclude: (a) fatty acylations of alpha z increase its association with membranes; (b) myristoylation is not required for palmitoylation of alpha z or for its productive interactions with adenylyl cyclase; (c) palmitoylation is not required for, but may instead inhibit, signaling by alpha z.

Potentiation of Gi-mediated phospholipase C activation by retinoic acid in HL-60 cells. Possible role of G gamma 2.

Differentiated HL-60 cells acquire responsiveness to fMet-Leu-Phe (fMLP), which activates phospholipase C and O2- generation in a pertussis toxin-sensitive manner. Addition of retinoic acid (RA) for the last 24 h during dimethyl sulfoxide (Me2SO)-induced differentiation enhanced fMLP-dependent signals and interaction between fMLP receptor and G(i). RA modifies both the function and subunit composition of G(i)2, the predominant G(i) of HL-60 membranes, as shown by comparing purified G(i)2 from membranes of Me2SO-treated cells (D-G(i)2) to G(i)2 from membranes of cells treated with both Me2SO and RA (DR-G(i)2). As compared to D-G(i)2, DR-G(i)2 induced more fMLP binding when added to membranes of pertussis toxin-treated HL-60 cells and, in the presence of GTP gamma S, stimulated beta gamma-sensitive phospholipase C in extracts of HL-60 cells to a much greater extent at a lower concentrations. Immunoblasts revealed that RA induced expression of the gamma 2 subunit, which was otherwise undetectable in G(i)2 purified from HL-60 cells or in HL-60 membranes. Possibly by inducing expression of gamma 2, RA alters two functions of the G(i) beta gamma subunit, modulation of fMLP receptor-G(i)2 coupling and activation of the effector, Phospholipase C.