RESUMO
BACKGROUND: Twenty-four-hour rhythmicity in mammalian tissues and organs is driven by local circadian oscillators, systemic factors, the central circadian pacemaker and light-dark cycles. At the physiological level, the neural and endocrine systems synchronise gene expression in peripheral tissues and organs to the 24-h-day cycle, and disruption of such regulation has been shown to lead to pathological conditions. Thus, monitoring rhythmicity in tissues/organs holds promise for circadian medicine; however, most tissues and organs are not easily accessible in humans and alternative approaches to quantify circadian rhythmicity are needed. We investigated the overlap between rhythmic transcripts in human blood and transcripts shown to be rhythmic in 64 tissues/organs of the baboon, how these rhythms are aligned with light-dark cycles and each other, and whether timing of tissue-specific rhythmicity can be predicted from a blood sample. RESULTS: We compared rhythmicity in transcriptomic time series collected from humans and baboons using set logic, circular cross-correlation, circular clustering, functional enrichment analyses, and least squares regression. Of the 759 orthologous genes that were rhythmic in human blood, 652 (86%) were also rhythmic in at least one baboon tissue and most of these genes were associated with basic processes such as transcription and protein homeostasis. In total, 109 (17%) of the 652 overlapping rhythmic genes were reported as rhythmic in only one baboon tissue or organ and several of these genes have tissue/organ-specific functions. The timing of human and baboon rhythmic transcripts displayed prominent 'night' and 'day' clusters, with genes in the dark cluster associated with translation. Alignment between baboon rhythmic transcriptomes and the overlapping human blood transcriptome was significantly closer when light onset, rather than midpoint of light, or end of light period, was used as phase reference point. The timing of overlapping human and baboon rhythmic transcriptomes was significantly correlated in 25 tissue/organs with an average earlier timing of 3.21 h (SD 2.47 h) in human blood. CONCLUSIONS: The human blood transcriptome contains sets of rhythmic genes that overlap with rhythmic genes of tissues/organs in baboon. The rhythmic sets vary across tissues/organs, but the timing of most rhythmic genes is similar in human blood and baboon tissues/organs. These results have implications for development of blood transcriptome-based biomarkers for circadian rhythmicity in tissues and organs.
Assuntos
Relógios Circadianos , Transcriptoma , Animais , Relógios Circadianos/genética , Ritmo Circadiano/genética , Humanos , Mamíferos/genética , Primatas/genéticaRESUMO
BACKGROUND: The genomic binding of CTCF is highly conserved across mammals, but the mechanisms that underlie its stability are poorly understood. One transcription factor known to functionally interact with CTCF in the context of X-chromosome inactivation is the ubiquitously expressed YY1. Because combinatorial transcription factor binding can contribute to the evolutionary stabilization of regulatory regions, we tested whether YY1 and CTCF co-binding could in part account for conservation of CTCF binding. RESULTS: Combined analysis of CTCF and YY1 binding in lymphoblastoid cell lines from seven primates, as well as in mouse and human livers, reveals extensive genome-wide co-localization specifically at evolutionarily stable CTCF-bound regions. CTCF-YY1 co-bound regions resemble regions bound by YY1 alone, as they enrich for active histone marks, RNA polymerase II and transcription factor binding. Although these highly conserved, transcriptionally active CTCF-YY1 co-bound regions are often promoter-proximal, gene-distal regions show similar molecular features. CONCLUSIONS: Our results reveal that these two ubiquitously expressed, multi-functional zinc-finger proteins collaborate in functionally active regions to stabilize one another's genome-wide binding across primate evolution.
Assuntos
Evolução Molecular , Primatas/genética , Proteínas Repressoras/metabolismo , Fator de Transcrição YY1/metabolismo , Animais , Fator de Ligação a CCCTC , Linhagem Celular , Genoma , Humanos , Camundongos , Proteínas Repressoras/químicaRESUMO
Copper chaperone for superoxide dismutase (CCS) is essential for transporting copper ion to Cu,Zn-superoxide dismutase (Cu,Zn-SOD). We cloned cDNAs for six primate species' CCSs. The total number of amino acid residues of primate CCSs is 274. Similarities between primates were over 96%. Important residues for the CCS function were well conserved. A phylogenetic tree of CCSs and Cu,Zn-SODs from various organisms showed that these two proteins were derived from a common ancestor, diverging very early on during eukaryote evolution. The high frequency of nonsynonymous substitutions was found in the lineage to Old World monkeys and apes. Expression of the CCS gene in various tissues of Japanese monkey was found to be high in the liver and adrenal gland, followed by the kidney and small intestine. Such expressional pattern was similar with that of Cu,Zn-SOD gene (Fukuhara et al., 2002).
Assuntos
Cobre/metabolismo , Evolução Molecular , Expressão Gênica , Chaperonas Moleculares/genética , Primatas/genética , Superóxido Dismutase/genética , Sequência de Aminoácidos , Animais , Clonagem Molecular , DNA Complementar/genética , Feminino , Masculino , Chaperonas Moleculares/metabolismo , Dados de Sequência Molecular , Filogenia , Análise de Sequência de DNA , Superóxido Dismutase/metabolismoAssuntos
Avaliação Pré-Clínica de Medicamentos , Isoenzimas , Modelos Animais , Preparações Farmacêuticas/metabolismo , Primatas , Animais , Carboxilesterase/genética , Carboxilesterase/metabolismo , Sistema Enzimático do Citocromo P-450/genética , Sistema Enzimático do Citocromo P-450/metabolismo , Glucuronosiltransferase/genética , Glucuronosiltransferase/metabolismo , Humanos , Preparações Farmacêuticas/administração & dosagem , Primatas/genética , Primatas/metabolismoRESUMO
Recently, we identified a susceptibility locus for human uric acid nephrolithiasis (UAN) on 10q21-q22 and demonstrated that a novel gene (ZNF365) included in this region produces through alternative splicing several transcripts coding for four protein isoforms. Mutation analysis showed that one of them (Talanin) is associated with UAN. We examined the evolutionary conservation of ZNF365 gene through a comparative genomic approach. Searching for mouse homologs of ZNF365 transcripts, we identified a highly conserved mouse ortholog of ZNF365A transcript, expressed specifically in brain. We did not found a mouse homolog for ZNF365D transcript encoding the Talanin protein, even if we were able to identify the corresponding genomic region in mouse and rat not yet organized in canonical gene structure suggesting that ZNF365D was originated after the branching of hominoid from rodent lineage. In mouse and in most mammals, a functional uricase degrades the uric acid to allantoin, but uricase activity was lost during the Miocene epoch in hominoids. Searching for the presence of Talanin in Primates, we found a canonical intron-exon structure with several stop codons preventing protein production in Old World and New World monkeys. In humans, we observe expression and we have evidence that ZNF365D transcript produces a functional protein. It seems therefore that ZNF365D transcript emerged during primate evolution from a noncoding genomic sequence that evolved in a standard gene structure and assumed its role in parallel with the disappearance of uricase, probably against a disadvantageous excessive hyperuricemia.
Assuntos
Proteínas de Ligação a DNA/genética , Nefropatias/genética , Fatores de Transcrição/genética , Processamento Alternativo , Sequência de Aminoácidos , Animais , Sequência de Bases , Cromossomos Humanos Par 10/genética , Cromossomos de Mamíferos/genética , Clonagem Molecular , DNA Complementar/química , DNA Complementar/genética , DNA Complementar/isolamento & purificação , Evolução Molecular , Humanos , Nefropatias/sangue , Nefropatias/patologia , Camundongos , Dados de Sequência Molecular , Filogenia , Primatas/genética , Isoformas de Proteínas/genética , Alinhamento de Sequência , Análise de Sequência de DNA , Homologia de Sequência de Aminoácidos , Homologia de Sequência do Ácido Nucleico , Sintenia , Ácido Úrico/sangue , Dedos de Zinco/genéticaRESUMO
Preclinical studies in nonhuman primates (NHP) play key roles in AIDS vaccine development efforts. In addition to their traditional utilization to gauge vaccine safety and immunogenicity, NHP models are currently employed to an unprecedented extent and in unprecedented ways in contemporary basic and applied vaccine development efforts. Current studies employ NHP models to probe fundamental mechanisms of primate immune system regulation, to investigate pathogenic mechanisms of AIDS, and to optimize immunization strategies involving novel vaccine vectors. The use of experimental challenges of immunized NHPs with either simian immunodeficiency virus or chimeric simian/human immunodeficiency virus to generate preclinical vaccine efficacy data has emerged as an important criterion for facilitating entry of a given vaccine candidate into early phase clinical evaluation in humans. However, for studies of the biology of AIDS virus transmission, AIDS virus disease pathogenesis and AIDS virus vaccine efficacy that are predicated on experimental viral challenge to be most valuable, additional efforts need to be devoted to generating challenge models that more closely recapitulate HIV-1 infection in humans. Towards this end, improved communication between clinical and preclinical investigators, to promote a bidirectional flow of information focusing on individual research needs and shared goals should enable the NHP models to most effectively expedite progress toward the development of a safe and effective AIDS vaccine.
Assuntos
Vacinas contra a AIDS/imunologia , Avaliação Pré-Clínica de Medicamentos , Primatas/imunologia , Vacinas contra a SAIDS/imunologia , Síndrome da Imunodeficiência Adquirida/imunologia , Síndrome da Imunodeficiência Adquirida/prevenção & controle , Síndrome da Imunodeficiência Adquirida/virologia , Animais , Modelos Animais de Doenças , HIV-1/genética , HIV-1/imunologia , HIV-2/imunologia , Humanos , Primatas/genética , Vírus Reordenados/imunologia , Síndrome de Imunodeficiência Adquirida dos Símios/imunologia , Síndrome de Imunodeficiência Adquirida dos Símios/prevenção & controle , Síndrome de Imunodeficiência Adquirida dos Símios/virologia , Vírus da Imunodeficiência Símia/genética , Vírus da Imunodeficiência Símia/imunologiaAssuntos
Pesquisa Biomédica/normas , Pesquisa Biomédica/tendências , Cooperação Internacional , Pesquisadores/normas , Academias e Institutos , Animais , Fibrilação Atrial/genética , Engenharia Biomédica/tendências , Pesquisa Biomédica/educação , Doenças Cardiovasculares/epidemiologia , China/epidemiologia , Indústria Farmacêutica/economia , Indústria Farmacêutica/tendências , Emigração e Imigração , Genética Médica/tendências , História Antiga , Humanos , Cooperação Internacional/história , Investimentos em Saúde , Modelos Animais , Seleção de Pessoal , Primatas/genética , Pesquisadores/educação , Recursos HumanosRESUMO
Mn- and Cu,Zn-superoxide dismutase (SOD) cDNAs of eight primate species, Pan troglodytes, Pongo pygmaeus, Hylobates lar, Macaca fuscata, Macaca fascicularis, Macaca mulatta, Cebus apella, and Callithrix jacchus, were cloned. The whole protein-coding sequences were covered, comparing 198 and 153 (or 154) amino acids, for Mn- and Cu,Zn-SODs, respectively. Residues forming metal ligands were completely conserved in the two primate SODs and nucleotide/amino acid substitutions were more frequent in Cu,Zn-SODs than in Mn-SODs. Molecular evolutionary analyses showed Mn-SOD to have evolved at a constant rate and its phylogenetic tree well reflected primate phylogeny. Cu,Zn-SOD was shown to have evolved differently between primate lineages. The significant high ratio of a non-synonymous/synonymous rate was found in the lineage leading to great apes and humans, showing that this lineage underwent positive Darwinian selection. Southern hybridization suggested that the genes for primate Mn- and Cu,Zn-SOD exist as single copies. Northern analysis in various Japanese monkey tissues showed Mn- and Cu,Zn-SOD expression to be high in the liver, kidneys, and adrenal glands.
Assuntos
Proteínas de Bactérias , Evolução Molecular , Primatas/genética , Superóxido Dismutase/genética , Sequência de Aminoácidos , Animais , Northern Blotting , Southern Blotting , Clonagem Molecular , DNA/genética , DNA/metabolismo , DNA Complementar/química , DNA Complementar/genética , Desoxirribonuclease EcoRI/metabolismo , Desoxirribonucleases de Sítio Específico do Tipo II/metabolismo , Feminino , Regulação Enzimológica da Expressão Gênica , Genes/genética , Masculino , Dados de Sequência Molecular , Filogenia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Alinhamento de Sequência , Análise de Sequência de DNA , Homologia de Sequência de AminoácidosRESUMO
Growth hormone (GH) evolution is very conservative among mammals, except for primates and ruminant artiodactyls. In fact, most known mammalian GH sequences differ from the inferred ancestral mammalian sequence by only a few amino acids. In contrast, the human GH sequence differs from the inferred ancestral sequence by 59 amino acids. However, it is not known when this rapid evolution of GH occurred during primate evolution or whether it was due to positive selection. Also, human growth hormone receptor (GHR) displays species specificity; i.e., it can interact only with human (or rhesus monkey) GH, not with nonprimate GHS: The species specificity of human GHR is largely due to the Leu-->Arg change at position 43, and it has been hypothesized that this change must have been preceded by the His-->Asp change at position 171 of GH. Is this hypothesis true? And when did these changes occur? To address the above issues, we sequenced GH and GHR genes in prosimians and simians. Our data supported the above hypothesis and revealed that the species specificity of human GHR actually emerged in the common ancestor of Old World primates, but the transitional phase still persists in New World monkeys. Our data showed that the rapid evolution of primate GH occurred during a relatively short period (in the common ancestor of higher primates) and that the rate of change was especially high at functionally important sites, suggesting positive selection. However, the nonsynonymous rate/synonymous rate ratio at these sites was <1, so relaxation of purifying selection might have played a role in the rapid evolution of the GH gene in simians, possibly as a result of multiple gene duplications. Similar to GH, GHR displayed an accelerated rate of evolution in primates. Our data revealed proportionally more amino acid replacements at the functionally important sites in both GH and GHR in simians but, surprisingly, showed few coincidental replacements of amino acids forming the same intermolecular contacts between the two proteins.
Assuntos
Evolução Molecular , Hormônio do Crescimento/genética , Primatas/genética , Receptores da Somatotropina/genética , Aminoácidos/genética , Animais , DNA/química , DNA/genética , DNA Complementar/química , DNA Complementar/genética , Variação Genética , Humanos , Dados de Sequência Molecular , Filogenia , Seleção Genética , Análise de Sequência de DNA , Especificidade da EspécieRESUMO
Ten new primate Mhc-DMB complete cDNA sequences have been obtained in chimpanzee (n=four), gorilla (n=three) and orangutan (n=three); this gene has not been previously studied in these species. Exonic allelism has been recorded all along the molecule domains and also in the leader peptide, but not in the transmembrane segment. An analysis of the residues critical in the conformation of the Mhc-DR peptide-binding site was done in order to look for a Mhc-DR homologue site; synonymous substitutions are favoured in this homologous HLA-DM region. This is another finding that supports the possibility that DM could not be typically presenting molecules. The immunoreceptor inhibition motif Tyr 230-Thr/Ser 231-Pro 232-Leu 233 (ITIM) is invariantly present in apes for at least 15 million years, and may have a double function: 1) To direct DMB-DMA molecules from the endoplasmic reticulum or cell surface towards the endosomal/lysosomal class II compartment and 2) to send an inhibitory signal to the cell in order to stop synthesis of unnecessary HLA-DR molecules, once all available antigenic peptides are loaded. Other molecules, like NK-cell receptors and Fc receptors, bear this type of tyrosine-based inhibitory motifs in order to switch off specific cell functions. DMB molecules (as previously shown in C4d molecules) do not present species-specific motifs in common chimpanzee, suggesting that this species is very close to gorilla or man; also, DMB, like C4d molecules, do not show a trans-species evolution pattern, suggesting the existence of extensive homogenization of DMB genes within each species or a recent generation of alleles. Finally, a clade grouping human and gorilla DMB cDNA sequences is obtained using a dendrogram (as for C4d trees); this is in contrast to others' results that obtain a human/chimpanzee clade using different DNA sequences.
Assuntos
Alelos , Genes MHC da Classe II , Variação Genética , Primatas/genética , Primatas/imunologia , Sequência de Aminoácidos , Animais , Sequência de Bases , Sequência Conservada , Primers do DNA/genética , DNA Complementar/genética , Evolução Molecular , Gorilla gorilla , Antígenos HLA-D/química , Antígenos HLA-D/genética , Antígenos de Histocompatibilidade Classe II/química , Antígenos de Histocompatibilidade Classe II/genética , Humanos , Dados de Sequência Molecular , Pan paniscus , Pan troglodytes , Pongo pygmaeus , Homologia de Sequência de Aminoácidos , Tirosina/genéticaRESUMO
There is an urgent need for a safe and effective vaccine to prevent human immunodeficiency virus (HIV) infection. Several HIV vaccine candidates have shown promise, but many concerns regarding the safety and efficacy of current vaccines remain. A major hindrance in HIV vaccine development is a poor understanding of precisely what functions HIV vaccines are required to perform in order to protect humans from HIV-1. Only higher primates (i.e. macaques, chimpanzees and humans) are susceptible to HIV-1 or the closely related virus 'simian immunodeficiency virus'. These species are outbred and there are remarkable genetic differences in both the immune responses to vaccines and their susceptibility to infection. The development of genetically identical macaques would be a major step towards dissecting what immune responses are required to protect from HIV infection. For example, live attenuated HIV-1 vaccines are likely to be highly efficacious, but will induce disease in a substantial proportion of recipients. Defining why a live attenuated vaccine is effective should allow safer vaccines to be developed, retaining only the immunologic properties of an effective vaccine. The reduction in 'background genetic noise' obtained by studying genetically identical primates would provide concise answers to critical HIV vaccine issues, by studying a minimal number of animals. Such an approach could potentially be employed in other diseases where non-human primates are the only available model. Small studies can be performed where identical twins are generated by embryo bisection; however, larger studies where multiple immune parameters are simultaneously evaluated would be facilitated by cloning technology. Despite the technical difficulties to be overcome, the potential gains in human health from the development of genetically identical non-human primates are worthy of careful consideration.
Assuntos
Vacinas contra a AIDS/farmacologia , Vacinas contra a AIDS/toxicidade , Primatas/genética , Primatas/imunologia , Animais , Clonagem de Organismos , Avaliação Pré-Clínica de Medicamentos , Infecções por HIV/genética , Infecções por HIV/imunologia , Infecções por HIV/prevenção & controle , HIV-1/imunologia , Humanos , Macaca/genética , Macaca/imunologia , Modelos BiológicosRESUMO
The major histocompatibility complex (MHC) class I region has been shown to be associated with a variety of immune and nonimmune disorders. In an effort to initiate steps designed to identify the idiopathic hemochromatosis disease gene (HFE), we have cloned and mapped two expressed messages using probes from the HLA-H subregion that lie immediately distal to the HLA-A9 breakpoint. Although the cDNA clones identify distinct multifragment families that are dispersed throughout the MHC, the gene sequences from which the two cDNA clones derive map centromeric to the HLA-B locus and are absent from the genomes of higher nonhuman primates. This suggests that a syntenic coding segment arose within a highly polymorphic region (TNF to HLA-B interval) as the result of an insertion event following the emergence of Homo sapiens. An additional syntenic cluster exists within a peak of linkage disequilibrium with the HFE gene and may define coding sequences that underlie the defect in genetic iron overload. These data generally support the concept that the class I region is potentially gene-rich and further highlight the possibility that these new coding sequences may play a role in the development of a variety of HLA-linked diseases. The observations presented suggest that interlocus exchanges have played a structural role in the genesis of the human class I region.
Assuntos
Genes MHC Classe I , Antígenos HLA-A/genética , Família Multigênica , Animais , Southern Blotting , Linhagem Celular , DNA Complementar/genética , Hemocromatose/genética , Humanos , Desequilíbrio de Ligação , Polimorfismo de Fragmento de Restrição , Primatas/genética , RNA Mensageiro/genética , Deleção de Sequência , Especificidade da EspécieRESUMO
Interleukin 11 (IL-11) is a multifunctional cytokine which may play a role in regulating the growth and development of cells in both the hematopoietic and lymphoid systems. IL-11 activity was originally detected in the conditioned medium of a primate bone marrow stromal cell line, and the human cDNA was cloned from a human fetal lung fibroblast cell line. The purified protein shows multifunctional activity, influencing lymphohematopoietic stem cell proliferation and differentiation, megakaryocyte progenitor cell proliferation and differentiation, erythroid progenitor cell proliferation, B lymphocyte maturation, activation of hepatocyte acute phase protein synthesis, and adipogenesis. At the molecular level, IL-11 is unique, containing no asparagine-linked glycosylation sites and no cysteine residues. The IL-11 receptor belongs to a family of cytokine receptors which includes the receptors for IL-6, leukemia inhibitory factor (LIF), oncostatin M (OSM), and ciliary neurotrophic factor (CNTF), which are all capable of interacting with the signal transducing receptor gp130 after ligand binding. IL-11 has demonstrated activity in preclinical models for the treatment of thrombocytopenia and, in some cases, neutropenia; studies are underway to confirm its usefulness in the clinic for treatment of myelosuppression associated with cancer chemotherapy and bone marrow transplantation.
Assuntos
Interleucina-11/fisiologia , Animais , Transplante de Medula Óssea , Diferenciação Celular/efeitos dos fármacos , Divisão Celular/efeitos dos fármacos , Avaliação Pré-Clínica de Medicamentos , Genes , Células-Tronco Hematopoéticas/efeitos dos fármacos , Humanos , Fatores Imunológicos/uso terapêutico , Interleucina-11/genética , Interleucina-11/farmacologia , Interleucina-11/uso terapêutico , Subunidade alfa de Receptor de Interleucina-11 , Interleucina-6/fisiologia , Tecido Linfoide/efeitos dos fármacos , Camundongos , Neutropenia/terapia , Primatas/genética , Receptores de Interleucina/genética , Receptores de Interleucina/metabolismo , Receptores de Interleucina-11 , Homologia de Sequência do Ácido Nucleico , Trombocitopenia/terapiaRESUMO
Nucleotide sequence analysis of the major noncoding region of human mitochondrial DNA from various races was extended with 72 Native Americans from 16 different local populations (nine populations from Chile, four from Colombia, and one each from Brazil and from Maya and Apache Indians). The sequences were determined directly from the polymerase chain reaction products. On the basis of a comparison of the 482-bp sequences in the 72 Native Americans, 43 different types of mitochondrial DNA sequences were observed. The nucleotide diversity within the Native Americans was estimated to be 1.29%, which is slightly less than the value of 1.44% from the total human population including Africans, Europeans, and Asians. Phylogenetic analysis revealed that most Native American lineages are classified into four major distinct clusters. Individuals belonging to each cluster share at least two specific polymorphic sites that are nearly absent in other human populations, indicating a unique phylogenetic position of Native Americans. A phylogenetic tree of 193 individuals including Africans, Europeans, Asians, and Native Americans indicated that the four Native American clusters are distinct and dispersed in the tree. These clusters almost exclusively consist of Native Americans--with only a few Asians, if any. We postulate that four ancestral populations gave rise to different waves of migration to the New World. From the estimated coalescence time of the Asian and Native American lineages, we infer that the first migration across the Bering landbridge took place approximately 14,000-21,000 years ago. Furthermore, sequence differences in all pairwise comparisons of Native Americans showed a bimodal distribution that is significantly different from Poisson. These results suggest that the ancestral Native American population underwent neither a severe bottleneck nor rapid expansion in population size, during the migration of people into the Americas.
Assuntos
Povo Asiático/genética , DNA Mitocondrial/genética , Indígenas Centro-Americanos/genética , Indígenas Norte-Americanos/genética , Indígenas Sul-Americanos/genética , Modelos Genéticos , Filogenia , América , Animais , Ásia/etnologia , Sequência de Bases , Emigração e Imigração , Frequência do Gene , Variação Genética , História Antiga , Humanos , Japão , Dados de Sequência Molecular , Reação em Cadeia da Polimerase , Polimorfismo Genético , Primatas/genética , Alinhamento de Sequência , Homologia de Sequência do Ácido NucleicoRESUMO
The measurements of Gigantopithecus blacki teeth from Liucheng are summarized and comparative statistics are calculated. Compared to the variability of tooth size in living nonhuman primates, emphasizing Pongo because of close ecological and genetic relationships, the hypothesis of a single species for Liucheng specimens is acceptable. The Liucheng sample pattern of variability in tooth size, as well as the South African robust australopithecines, differs from living nonhuman primates in the same way; the first molar is not the least variable of the molars, and the molars are relatively variable compared with the other teeth. Sexual dimorphism may be responsible for this pattern in both cases. Later G. blacki cheek teeth are significantly larger than early G. blacki. From early G. blacki to later, the expansion ratio in tooth breadth is more than in tooth length. The pattern of expansion is like that seen in South African australopithecines. Both G. blacki and South African robust Australopithecus seem to show continued adaptation to more powerful and efficient chewing. The diet of G. blacki is not known, and its reconstruction depends on estimates of body size that differ greatly.