Non-primate Growth Hormones Research Articles

Different intracellular signalling properties induced by human and porcine growth hormone

Growth hormone (GH) is reportedly species-specific. Primate growth hormone can trigger non-primate growth hormone receptor (GHR), but primates GHR cannot be activated by non-primate GH. However, it is also unclear that why primate GH and non-primate GH have different biological activities. Thus, we analysed primate growth hormone (human growth hormone (hGH)) or non-primate GH (porcine growth hormone (pGH))-induced intracellular signalling in 3T3-F442A cells and rat hepatocytes in a dose- and time-dependent manner to explore the different biological activities between them. The results revealed that both hGH and pGH can activate Janus kinase 2 (JAK2), Signal transducers and activators of transcription 1, 3 and 5 (STATs 1, 3 and 5) and extracellular signal-regulated kinase 1/2 (ERK1/2). There were no significant differences in JAK2 or ERK1/2 tyrosine phosphorylation after hGH and pGH treatment, but there were different between hGH and pGH in STAT/1/3/5 tyrosine phosphorylation, and JAK2, STAT/1/3/5 tyrosine phosphorylation was time-dependent and dose-dependent, whereas ERK1/2 was not. Both hGH and pGH demonstrated similar kinetics for STATs 1, 3 and 5 phosphorylation, but the pGH-mediated tyrosine phosphorylation was weaker than that mediated by hGH. Our observations indicated that the levels of main signalling proteins phosphorylation triggered by hGH or pGH were not exactly the same, which may explain the different biological activities showed by primate GH and non-primate GH.

Transgenic mice expressing the human growth hormone gene provide a model system to study human growth hormone synthesis and secretion in non-tumor-derived pituitary cells: Differential effects of dexamethasone and thyroid hormone

Growth hormone (GH) is regulated by pituitary and hypothalamic factors as well as peripheral endocrine factors including glucocorticoids and thyroid hormone. Studies on human GH are limited largely to the assessment of plasma levels in endocrine disorders. Thus, insight into the regulation of synthesis versus secretion has come mainly from studies done on non-human GH and/or pituitary tumor cells. However, primate and non-primate GH gene loci have differences in their structure and, by extension, regulation. We generated transgenic (171hGH/CS-TG) mice containing the intact hGH1 gene and locus control region, including sequences required for integration-independent and preferential pituitary expression. Here, we show hGH co-localizes with mouse (m) GH in somatotrophs in situ and in primary pituitary cells. Dexamethasone treatment increased hGH and mGH, as well as GH releasing hormone (GHRH) receptor RNA levels, and hGH release was stimulated by GHRH treatment. By contrast, triiodothyronine decreased or had no effect on hGH and mGH production, respectively, and the negative effect on hGH was also seen in the presence of dexamethasone. Thus, 171hGH/CS-TG mouse pituitary cultures represent a model system to investigate hormonal control of hGH synthesis and secretion.

Functional promiscuity of squirrel monkey growth hormone receptor toward both primate and nonprimate growth hormones.

Primate growth hormone (GH) has evolved rapidly, having undergone approximately 30% amino acid substitutions from the inferred ancestral eutherian sequence. Nevertheless, human growth hormone (hGH) is physiologically effective when administered to nonprimate mammals. In contrast, its functional counterpart, the human growth hormone receptor (hGHR), has evolved species specificity so that it responds only to Old World primate GHs. It has been proposed that this species specificity of the hGHR is largely caused by the Leu --> Arg change at position 43 after a prior His --> Asp change at position 171 of the GH. Sequence analyses supported this hypothesis and revealed that the transitional phase in the GH:GHR coevolution still persists in New World monkeys. For example, although the GH of the squirrel monkey has the His --> Asp substitution at position 171, residue 43 of its GHR is a Leu, the nonprimate residue. If the squirrel monkey truly represents an intermediate stage of GH:GHR coevolution, its GHR should respond to both hGH and nonprimate GH. Also, if the emergence of species specificity was a result of the selection for a more efficient GH:GHR interaction, then changing residue 43 of the squirrel monkey growth hormone receptor (smGHR) to Arg should increase its binding affinity toward higher primate GH. To test these hypotheses, we performed protein-binding assays between the smGHR and both human and rat GHs, using the surface plasmon resonance methodology. Furthermore, the effects of reciprocal mutations at position 43 of human and squirrel monkey GHRs are measured for their binding affinities toward human and squirrel monkey GHs. The results from the binding kinetic assays clearly demonstrate that the smGHR is in the intermediate state of the evolution of species specificity. Interestingly, the altered residue Arg at position 43 of the smGHR does not lead to an increased binding affinity. The implications of these results on the evolution of the GH:GHR interaction and on functional evolution are discussed.

Cloning and characterisation of the GH gene from the common dolphin ( Delphinus delphis)

The sequence of growth hormone (GH) is generally strongly conserved in mammals, but episodes of rapid change occurred during the evolution of primates and artiodactyls, when the rate of GH evolution apparently increased at least 50-fold. As a result, the sequences of human and ruminant GHs differ substantially from those of other non-primate GHs. Recent molecular studies have suggested that cetaceans are closely related to artiodactyls and may be deeply nested within the artiodactyl phylogenetic tree. To extend the knowledge of GH in Cetartiodactyla (Artiodactyla plus Cetacea), we have cloned and characterised a single GH gene from the common dolphin ( Delphinus delphis), using genomic DNA and a polymerase chain reaction technique. As in other mammals, the dolphin GH gene comprises five exons and four introns. The deduced sequence for the mature dolphin GH differs from that of pig at two residues only, showing that the apparent burst of rapid evolution of GH occurred largely after the separation of cetaceans and ruminants.

The species specificity of growth hormone requires the cooperative interaction of two motifs

Primate growth hormones (GH) activate both primate and non-primate somatotrophic receptors (GH receptors), but non-primate GHs do not activate primate GH receptors. Previous studies argued the interaction of Asp171 of human GH and Arg43 of the receptor produced an attractive ionic interaction. In non-primate GHs, His170 replaces the homologous Asp171, producing a repulsive interaction with Arg43 of the primate receptor which was believed to reduce the attraction of non-primate GH for the human GH receptor, thus providing species specificity. In this report, H170D bovine GH had activity and affinity for human GH receptors approaching those of human GH. In contrast, replacing Asp171 of human GH with His did not significantly reduce somatotrophic activity, indicating that species specificity is not wholly explained by this residue’s interaction with Arg43 of the receptor. Deletion of either Phe44 (a residue present only in primate GHs) or residues 32–46 (20-kDa form of human GH) each only marginally reduced somatotrophic activities. But the combination of the D171H mutation with either ΔPhe44 or Δ32–46 in human GH reduced binding and activity in a greater than additive fashion, indicated a functional interaction between these distant structural features. In bovine GH addition of phenylalanine at position 44 increased the somatotrophic activity and receptor affinity in cells containing the human GH receptor. The combination of the H170D mutation and the addition of phenylalanine at position 44 created a bovine GH with activity indistinguishable from wild-type human GH. Based on evidence from both bovine and human GHs, the cooperative interaction of these two distant motifs determined the species specificity and indicated that structural plasticity was a critical feature necessary for the species specificity of somatotrophic activity.

A single arginine residue determines species specificity of the human growth hormone receptor.

Although growth hormone (GH) receptors (GHRs) in many species bind human (h) GH as well as their own GH, the hGHR only binds primate GH. Arg43 in hGHR interacts with Asp171 of hGH. Nonprimates have a His in the position equivalent to residue 171 of primate GH and a Leu in position 43 of primate GHR. To determine whether Arg43 accounts for the species specificity of the hGHR, point mutations that changed Leu43 to Arg were introduced into the cDNAs encoding the bovine (b) GHR or the rat GH binding protein (GHBP) and these mutants or their wild-type (WT) counterparts were expressed in mouse L cells. Binding of hGH or bGH to transfected cells or to GHBP secreted into the incubation medium was assessed by displacement of 125I-labeled hGH. WT and mutant bGHR bound hGH with similar affinity, but the affinity of the mutant receptors for bGH was reduced 200-fold. Likewise, WT and mutant GHBP bound hGH with equal affinity, but only WT GHBP bound bGH. Cross-linking of 125I-labeled hGH to WT or mutant GHR produced a 141-kDa labeled complex whose appearance was blocked by unlabeled hGH, but bGH blocked cross-linking only to WT receptors. Both hGH and bGH stimulated tyrosine phosphorylation of a 95-kDa protein in cells transfected with WT GHR, but bGH was less effective in cells expressing mutant GHR. We conclude that incompatibility of Arg43 in the hGHR with His171 in nonprimate GH is the major determinant of species specificity.

Involvement of zinc in binding of hGH to the prolactin receptor

This chapter describes the species specificity and structure–function relationships of growth hormone (GH). GH is a protein hormone derived from the anterior pituitary gland. It comprises a single polypeptide chain of about 190 amino acids, containing two intrachain disulfide bridges. The apparent rapid evolution of GH in the primates makes sequence studies on GH for primates other than man of particular interest. The sequence of GH from the rhesus monkey has been reported. This is very similar to that of human GH, differing at only four residues, suggesting that the rate or evolution of GH because the divergence of old-world monkeys and great apes has been slow. The implication is that GH underwent a period of remarkably rapid evolution early in primate evolution. An alternative explanation is that the primate and nonprimate GH genes are not strictly homologous, implying that a second gene is present in the human genome, homologous with nonprimate GHs and a second gene is present in the nonprimate mammalian genome, homologous with human GH. The situation is complicated by the occurrence of a cluster of GH-like genes in man.

Solubilization and characterization of a lactogenic receptor from human placental chorion membranes

Prolactin has a wide range of actions, including osmoregulation and the control of mammary gland development and lactation. These effects are mediated through a high-affinity cell surface receptor, which has been well characterized in a number of animal tissues. The molecular characteristics of the human receptor are unknown, however. The present studies were initiated, therefore, to determine the binding and molecular characteristics of the lactogenic receptor of human placental chorion membranes. Subcellular fractionation studies showed that the bulk of the receptor sedimented in the microsomal fraction at 45,000gav. Endogenous ligand was dissociated from the receptor with 3.5 M MgCl2 or 0.05 M acetate buffer (pH 4.8) with preservation of binding activity. The microsomal receptor bound human growth hormone (hGH), human prolactin (hPRL), ovine prolactin (oPRL), and human placental lactogen (hPL) but not non-primate growth hormones, indicating a narrow specificity for lactogenic hormones. The binding was only partially reversible in agreement with the known binding kinetics of animal lactogenic receptors. The receptor was solubilized with 45% yield from the microsomes using 16 mM 3-[(3-cholamidopropyl)dimethylammonio]-1-propane sulphonate (CHAPS) detergent-250 mM NaCl, and the binding activity was fully restored by a two-fold dilution in the binding reaction to reveal a KD of 0.8 nM for hGH and a binding capacity of 200 fmol of specifically bound hGH per mg of microsomal protein. Gel filtration chromatography indicated the minimum molecular weight of the ligand-receptor complex was approximately 60,000 daltons, and sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) of covalently cross-linked 125I-hGH-receptor complexes revealed a molecular size of 58,000 daltons. When account was taken of the contribution of the ligand, a molecular weight of 36,000 for the receptor's binding domain was obtained. These data indicate that the chorion lactogenic receptor has very similar binding and molecular characteristics to the lactogenic receptors from other mammalian species. Chorion membranes are thus a convenient source of material for the further purification and characterization of the human lactogenic receptor.

Conformational comparison in the growth hormone family

1. 1. The method of Kubota et al. [ Biochim. biophys. Acta 701, 242–252 (1982)] was applied to several members of the growth hormone family in order to examine their conformational homology. 2. 2. The method neither detects differences between rat, cow, sheep, horse and alpaca hormones, nor between monkey and human hormones. 3. 3. Lack of homology between primate and non-primate growth hormones was found in segments 42–49 and 184–191. The first fragment could be linked to species-specificity.

An electrostatic model for the interaction between growth hormone and its receptor involving chelation of Ca 2+ to the human growth hormone molecule

We previously postulated the local involvement of cations in the complex between human growth hormone and its receptors in the liver. The original electrostatic model involved convergence of unit negative charges on the hormone and receptor, towards an interposed Ca2+ ion. That model was consistent with (a) the Ca2+ dependence of human growth hormone binding, (b) the magnitude of the Ca2+ mediated increase in the affinity of human growth hormone binding, and (c) could also explain the relative affinities of human and non-primate growth hormones for growth hormone receptors. In the present report, the original electrostatic model is revised with the postulate that Ca2+ is chelated to the human growth hormone molecule. The consequences of this postulate are explored mathematically with the result that it becomes necessary to propose an additional unique hindrance determinant (positive residues in helices one and four are good candidates) to account for the lower affinity of non-primate growth hormones relative to human growth hormone. Predictions are made regarding the effect of a particular point mutation (at position 34) on the affinity of hormone binding.

Human growth hormone stimulates somatomedin C/insulin-like growth factor I production by the human lymphoid cell line, IM-9

The human lymphoid cell line, IM-9, is known to possess receptors for human growth hormone (hGH), but the only biological response that has been shown to follow binding of this hormone to the cells is receptor down-regulation. We have studied the actions of hGH on production of insulin-like growth factor I (IGF-I) by IM-9 cells. In order to demonstrate effects cells had to be transferred to a serum-free medium in which cell multiplication almost ceased, and cell viability fell to 50–60%. hGH stimulated IGF-I production by up to 400%. The effect was dose-related, but the dose-response curve was bimodal, with peaks of activity at approximately 15 ng/ml and 1000 ng/ml hGH. The effect of hGH was of slow onset, becoming significant only after about 24 h, and approaching a maximum after 2–5 days of treatment. hGH had a much greater stimulatory effect than non-primate growth hormones. The physiological significance of the effect observed is not yet clear, but it is apparent that the IM-9 line is a potentially useful model for study of the actions of growth hormone.

Diagnosis and treatment of children with growth hormone deficiency

Growth hormone deficiency embraces a heterogeneous group of disorders with multiple aetiologies. The biggest single division is between those children whose growth hormone deficiency is due to a structural lesion and those where it is not. In both cases other pituitary hormones may, or may not, be affected, although when a structural lesion is present multiple pituitary hormone deficiency is the rule rather than the exception. In the absence of structural lesions the pathogenesis of the condition in still largely obscure, although some mechanisms, such as cranial irradiation, are now well recognized. Birth trauma is also now a strong candidate as a predisposing factor. The prevalence of growth hormone deficiency is still uncertain, but is probably about 1 in 4000 live births. The clinical features of growth hormone deficiency are usually fairly clear, with short stature, low growth velocity, excess subcutaneous fat and delayed skeletal maturation being the principal clinical features. Laboratory investigation still largely depends upon the assessment of pituitary growth hormone secretion in response to a variety of provocation tests and is still in many ways unsatisfactory. Treatment consists of parenteral growth hormone replacement using material of human cadaveric origin. Non-primate growth hormones are of no value. Other endocrine abnormalities, when present, are treated appropriately, and with early diagnosis and optimal therapy the height prognosis is reasonably good. The principal aims for the future must be to ensure supplies of therapeutic growth hormone, improve some of the diagnostic procedures and ensure early ascertainment.

Purification and partial characterization of a nonprimate growth hormone receptor.

Pregnant rabbit liver membranes have been shown to possess two types of receptors by displacement analysis, a growth hormone (GH) receptor which binds bovine growth hormone with an affinity constant (KA) of 3 x 10(9) M-1 and ovine prolactin with a KA of 3 x 10(8) M-1, and a prolactin (Prl)-specific receptor which binds ovine prolactin with a KA of 5 x 10(9) M-1. The prolactin-specific receptor when solubilized with Triton exhibits a 4-fold increase in the its KA while the KA of the growth hormone receptor decreases slightly to 2 x 10(9) M-1 after solubilization. The 10-fold difference in affinity which results has been exploited to facilitate the separation of these two receptors by differential affinity chromatography on human growth hormone (hGH) affinity gels. The growth hormone receptor is eluted from the gel with 4 M urea while 5 M MgCl2 is required to elute the prolactin receptor. Conditions of affinity chromatography have been optimized, and further purification of the GH receptor by preparative isoelectric focusing and Sepharose 6B gel filtration resulted in a more than 8000-fold purification of the receptor. This material had a Stokes radius of 62 A, consistent with a molecular weight of 300,000 and gave one main band (75,000 to 80,000) and two minor bands on sodium dodecyl sulfate (SDS) polyacrylamide gels, which could be interpreted as indicating a tetrameric receptor. The GH receptor was shown to be a sialoglycoprotein (or closely associated with sialoglycoprotein) by analytical isoelectric focusing with an isoelectric point of 4.6. Specificity studies with the highly purified receptor confirmed the initial hypothesis that this receptor is capable of binding bovine growth hormone (bGH) with high affinity and ovine prolactin (oPrl) with low affinity, in contrast to the prolactin-specific receptor.

Reactivity of Non-Primate Growth Hormones and Prolactins with Human Growth Hormone Receptors on Cultured Human Lymphocytes

Ovine placental lactogen is as reactive as human growth hormone with the human growth hormone receptor of cultured human (IM-9) lymphocytes, which confirms the findings of Carr and Friesen with receptors of human liver. We now also show that bovine and ovine growth hormones and ovine prolactin have reactivity for the human growth hormone receptor on IM-9 lymphocytes that is of the same order of magnitude (0.03%) as that previously reported for human placental lactogen. The binding studies predict that these non-primate hormones will have biological effects on skeletal growth in primates, either as agonists or antagonists. Previous studies have shown that when IM-9 lymphocytes are exposed to human growth hormone for 18 h at 37 C, there is a time and concentration dependent loss of human growth hormone receptors, and the magnitude of the loss of receptors after preincubation for 18 h at 37 C is greater than the average occupancy of receptors under steady state conditions for 90 min at 30 C. In the present study we show that human and ovine placental lactogens, ovine prolactin, and bovine and ovine growth hormones also produce this effect on the human growth hormone receptor. Since the cellular process by which a hormone induces loss of its own receptors appears to require binding of the hormone to its receptor as well as one or more subsequent steps in hormone action, it is likely that all of the preparations that induce receptor loss will be shown to have some agonist activity of human growth hormone in promoting skeletal growth in primates. Further, these studies extend the interrelationships between primate and non-primate pituitary and placental hormones from what has been suggested previously from biological and structural studies.

Characterization of the binding of human growth hormone to microsomal membranes from rat liver.

The binding of 125I-labelled human growth hormone to the 100000g microsomal membrane fraction prepared from the livers of normal female rats was dependent on time, temperature, pH, membrane concentration and concentration of 125I-labelled human growth hormone. At 22 degrees C binding reached a steady state after 16h, with the mean maximal specific binding being 20% of the tracer initially added. Dissociation of 125I-labelled human growth hormone from the membranes, after addition of excess of unlabelled hormone, was relatively slow with a half-time greater than 24h. Only minor degradation of the 125I-labelled human growth hormone was observed during incubation with membranes for 16 or 25h at 22 degrees C. Similarly, no significant change in the ability of membranes to bind human growth hormone was evident after preincubation of the membranes for 16 or 25h. Specificity studies showed that up to 90% of the 125I-labelled human growth hormone bound could be displaced by 1 mug of unlabelled hormone. Ovine prolactin also showed considerable competition for the binding site. Non-primate growth-hormone preparations (ovine, bovine, porcine and rat) and non-related hormones (insulin, thyrotropin, lutropin and follitropin) all showed negligible competition. Scatchard analysis of the binding data was consistent with two classes of binding site with binding affinities of 0.64 X 10(10) +/- 0.2 X 10(10)M-1 and 0.03 X 10(10) +/- 0.007 X 10(10)M-1 and corresponding binding capacities of 98.4 +/- 10 fmol/mg of protein and 314.6 +/- 46.3 fmol/mg of protein. These studies provide data which, in general, are consistent with the criteria required for hormone-receptor interaction. However, proof of the thesis that the human-growth-hormone-binding sites in female rat liver represent physiological receptors must await the demonstration of a correlation between hormone binding and a biological response.

Assay of Lactogenic Hormones Using Receptors Isolated from Rabbit Liver

Using 125-I-labelled ovine prolactin and receptors isolated from the livers of rabbits, a sensitive method has been developed suitable for the assay of ovine, bovine, porcine, human and rat prolactins. These hormones showed competitive displacement of 125-I-ovine prolactin which was in general agreement with their respective activities in the pigeon crop sac bioassay. Human and monkey growth hormones and human placental lactogen, which have marked prolactin-like actions on mammary tissue were also effective competitors. Non-primate growth hormones (ovine, bovine, equine and canine) which do not have prolactin-like activity gave little if any displacement as did human FSH, LH, TSH, ACTH and bovine insulin. Preparations of equine and canine prolactin of varying purity gave dose-response curves although their activity as competitors relative to ovine prolactin was poor and not related to their pigeon crop stimulating activity. This indicates species differences between prolactins in hormone-receptor interaction. Experiments with antiserum to human growth hormone have suggested an effective method of making the assay specific in species such as man in which prolactin is not the sole hormone with lactogenic activity.

Binding of 125I-Human Growth Hormone to Specific Receptors in Human Cultured Lymphocytes: CHARACTERIZATION OF THE INTERACTION AND A SENSITIVE RADIORECEPTOR ASSAY

Abstract The interaction of human growth hormone with human lymphocytes from an established culture (IM-9) was studied using 125I-human growth hormone. The binding of 125I-human growth hormone was rapid; with human growth hormone at 0.1 nm a steady state was observed in 90 min at 30°. Bound labeled human growth hormone was dissociated rapidly by addition of excess unlabeled human growth hormone. Binding of 125I-human growth hormone to cultured lymphocytes was relatively insensitive to alterations in the pH and in the concentrations of Ca2+, Mg2+, or EDTA. At 30° there was very little degradation of labeled human growth hormone or of the specific receptor sites. Tryptic digestion destroyed the capacity of cells to bind human growth hormone. The IM-9 cells bound all human growth hormone preparations but not unrelated hormones or nonprimate growth hormones. The binding of 125I-human growth hormone was inhibited 10 to 14% with 1 to 2 ng per ml of unlabeled human growth hormone and 50% with 30 to 40 ng per ml, well within the range of hormone concentrations in vivo. Analysis of steady state data revealed a single order of binding sites with an affinity constant of 1.3 x 109 m-1 and about 4000 binding sites per cell. Numerous human growth hormone preparations were assayed by use of this receptor system as well as by immunoassay and by bioassay in vivo. The potencies of the human growth hormone preparations determined by the radioreceptor assay correlated more closely with the bioassay than with the radioimmunoassay.

Metabolic effects of plasmin digests of human growth hormone in the rat and man.

As a first step in our study of structure-function relationships among primate and non-primate growth hormones, human growth hormone (hGH) was subjected to the limited digestive activity of human plasmin. The lyophilized whole digest, containing less than 2% of unchanged hormone, had an average of 2.3 new amino-terminal groups per mole. The digest had the same potency as the native hormone (a) in causing weight gain in hypophysectomized rats; (b) in stimulating somatomedin production in hypophysectomized rats; (c) in stimulating upake of [(3)H]leucine into isolated diaphragm of hypophysectomized rats; (d) in accelerating transport of [(14)C]alpha-aminoisobutyric acid into isolated diaphragm of hypophysectomized rats; (e) in stimulating uptake of [3-0-methyl-(14)C]glucose by isolated adipose tissue of hypophysectomized rats; (f) in accelerating conversion of [(14)C]glucose to (14)CO(2) by isolated epididymal adipose tissue of hypophysectomized rats. The digest also caused glucosuria in partially pancreatectomized rats treated with dexamethasone. These metabolic actions of plasmin-digested hGH in the array of animal tests were confirmed by comparable effects elicited in 11 human subjects (nine pituitary-deficient children and adolescents and two nondeficient adults). A single injection of the plasmin digest caused an increase in plasma free fatty acids and a fall in plasma amino acids. Seven daily injections caused positive balances of nitrogen, phosphorous, sodium, and potassium, gain in body weight, and in two of three subjects impairment of glucose tolerance. The potency of the plasmin digest in producing these metabolic effects in man was comparable to that of native hGH.Thus, 2-3 bonds in the hGH molecule can be cleaved by plasmin without impairing the hormone's growthpromoting, anabolic, diabetogenic, and adipokinetic actions for rat and man.

Immunochemical relatedness of porcine, bovine, ovine and primate pituitary growth hormones.

Quantitative micro-complement fixation with rabbit antisera to chromatographicaUy purified human growth hormone has been used to measure immunologically the relatedness of 12 nonhuman primate pituitary growth hormones to human growth hormone. Using complement-fixation curves obtained with antihuman growth hormone, the degree of relatedness of nonhuman primate growth hormone to human growth hormone was estimated from the ratio, at antigen-antibody equivalence, of the complement fixed with the cross-reacting (nonhuman) antigen to the complement fixed with the homologous (human) antigen. The reproducibility of this method of estimating the extent of immunological cross-reactions was indicated by a standard error of the mean value for 3 replicated assays of ±5%. Significant differences in immunological activity between growth hormones within the order of primates were found. The degree of relatedness, measured immunochemically, of nonhuman primate growth hormones to human growth hormone appeared to be related to primate phylogeny based on nonmolecular evidence. Significant immunological similarity between 3 nonprimate growth hormones and human growth hormone was detected by direct complement fixation with porcine, bovine and ovine growth hormones and antihuman growth hormone. Higher concentrations of antihuman growth hormone were required to measure directly the cross-reactions than were used for measuring the homologous, human antigen. However, the complement fixation obtained with human growth hormone and its own antiserum at homologous antibody concentrations was inhibited by the nonprimate growth hormones. Porcine growth hormone fixed more complement with antihuman growth hormone than bovine or ovine growth hormones. The direct fixation of complement with antihuman growth hormone and bovine growth hormone was significantly improved by reduction of 2 of the 4 disulfide bonds in the bovine hormone. We conclude that some of the antigenic determinants of human growth hormone and of porcine, bovine and ovine growth hormones are similar if not identical.(Endocrinology77: 563,1965)