Local Anesthetic Bupivacaine Inhibits Melanoma Proliferation and Metastasis by Targeting PAPSS2.

BACKGROUND: Melanoma is a highly invasive skin cancer with limited treatment strategies. Bupivacaine, a commonly used local anesthetic recognized for its safety, has shown promise in combating tumors. 3'-phosphoadenosine 5'-phosphosulfate synthase 2 (PAPSS2) is a key enzyme in the sulfation process and is associated with the development and metastasis of various tumors. This study aimed to explore the mechanism by which bupivacaine inhibits melanoma proliferation and metastasis by targeting PAPSS2. METHODS: The effects of bupivacaine on the proliferation of A375 and A2058 melanoma cells were evaluated using Cell Counting Kit-8 (CCK-8), 5-Ethynyl-2'-deoxyuridine (EdU) labeling, and clonogenic assays. Cell migration, invasion, and PAPSS2 expression were evaluated using Transwell experiments and Quantitative Reverse Transcription Polymerase Chain Reaction (qRT-PCR) analysis. Additionally, an in vivo melanoma tumor model in nude mice was constructed to evaluate the impact of bupivacaine on melanoma growth and metastasis. Immunohistochemistry was used to assess tumor metastasis and PAPSS2 expression levels in the nude mouse model. RESULTS: Experimental results demonstrated that bupivacaine significantly inhibited melanoma proliferation and invasion compared to the control group. Notably, this inhibitory effect was partially reversed by PAPSS2 overexpression. In vivo experiments demonstrated that bupivacaine-treated nude mice exhibited reduced tumor volumes, weights, and fewer lung metastatic foci. Molecular analysis via qRT-PCR and immunohistochemistry analysis further indicated that bupivacaine significantly reduced PAPSS2 in tumor tissues. CONCLUSION: This study confirms that bupivacaine, a local anesthetic, can inhibit melanoma proliferation and metastasis by targeting the PAPSS2 signaling pathway. These findings suggest its potential as an anti-tumor medication and present new treatment strategies for melanoma.

Sulfation of chondroitin and bile acids converges to antagonize Wnt/ß-catenin signaling and inhibit APC deficiency-induced gut tumorigenesis.

Sulfation is a crucial and prevalent conjugation reaction involved in cellular processes and mammalian physiology. 3'-Phosphoadenosine 5'-phosphosulfate (PAPS) synthase 2 (PAPSS2) is the primary enzyme to generate the universal sulfonate donor PAPS. The involvement of PAPSS2-mediated sulfation in adenomatous polyposis coli (APC) mutation-promoted colonic carcinogenesis has not been reported. Here, we showed that the expression of PAPSS2 was decreased in human colon tumors along with cancer stages, and the lower expression of PAPSS2 was correlated with poor prognosis in advanced colon cancer. Gut epithelial-specific heterozygous Apc deficient and Papss2-knockout (ApcΔgut-HetPapss2Δgut) mice were created, and the phenotypes were compared to the spontaneous intestinal tumorigenesis of ApcΔgut-Het mice. ApcΔgut-HetPapss2Δgut mice were more sensitive to gut tumorigenesis, which was mechanistically accounted for by the activation of Wnt/ß-catenin signaling pathway due to the suppression of chondroitin sulfation and inhibition of the farnesoid X receptor (FXR)-transducin-like enhancer of split 3 (TLE3) gene regulatory axis. Chondroitin sulfate supplementation in ApcΔgut-HetPapss2Δgut mice alleviated intestinal tumorigenesis. In summary, we have uncovered the protective role of PAPSS2-mediated chondroitin sulfation and bile acids-FXR-TLE3 activation in the prevention of gut carcinogenesis via the antagonization of Wnt/ß-catenin signaling. Chondroitin sulfate may be explored as a therapeutic agent for Papss2 deficiency-associated colonic carcinogenesis.

Integrative high-throughput enhancer surveying and functional verification divulges a YY2-condensed regulatory axis conferring risk for osteoporosis.

The precise roles of chromatin organization at osteoporosis risk loci remain largely elusive. Here, we combined chromatin interaction conformation (Hi-C) profiling and self-transcribing active regulatory region sequencing (STARR-seq) to qualify enhancer activities of prioritized osteoporosis-associated single-nucleotide polymorphisms (SNPs). We identified 319 SNPs with biased allelic enhancer activity effect (baaSNPs) that linked to hundreds of candidate target genes through chromatin interactions across 146 loci. Functional characterizations revealed active epigenetic enrichment for baaSNPs and prevailing osteoporosis-relevant regulatory roles for their chromatin interaction genes. Further motif enrichment and network mapping prioritized several putative, key transcription factors (TFs) controlling osteoporosis binding to baaSNPs. Specifically, we selected one top-ranked TF and deciphered that an intronic baaSNP (rs11202530) could allele-preferentially bind to YY2 to augment PAPSS2 expression through chromatin interactions and promote osteoblast differentiation. Our results underline the roles of TF-mediated enhancer-promoter contacts for osteoporosis, which may help to better understand the intricate molecular regulatory mechanisms underlying osteoporosis risk loci.

Bone Phenotype is Always Present But Androgen Excess is Less Frequently Seen in PAPSS2 Deficiency

3'-Phosphoadenosine 5'-phosphosulfate synthase 2 (PAPSS2) deficiency is a rare disorder due to biallelic pathogenic variants in the PAPSS2 gene. This disorder was first described in 1998 by Ahmad et al. and Faiyaz ul Haque et al. To date, 79 patients with PAPSS2 deficiency have been reported. The main reported features of these patients are related to bone abnormalities and clinical/biochemical androgen excess. Disproportionate short stature and symptoms associated with spondylar skeletal dysplasia are the most common clinical features that require clinical attention. Androgen excess has been described much less commonly. This review summarizes the currently published clinical, molecular, and biochemical features of patients with PAPSS2 deficiency.

Clinical Diagnostic Values of Transfer RNA-Derived Fragment tRF-41-YDLBRY73W0K5KKOVD and its Effects on the Growth of Gastric Cancer Cells.

Gastric cancer (GC) is a serious disease with high mortality and poor prognosis. It is known that tRNA halves play key roles in the progression of cancer. This study explored the function of the tRNA half tRF-41-YDLBRY73W0K5KKOVD in GC. Quantitative real-time reverse transcription-polymerase chain reaction was used to measure RNA levels. The level of tRF-41-YDLBRY73W0K5KKOVD in GC cells was regulated by its mimics or inhibitor. Cell proliferation was evaluated by using a Cell Counting Kit-8 and EdU cell proliferation assay. A Transwell assay was used to detect cell migration. Flow cytometry was used to measure cell cycle and apoptosis. The results showed that tRF-41-YDLBRY73W0K5KKOVD expression was decreased in GC cells and tissues. Functionally, overexpression of tRF-41-YDLBRY73W0K5KKOVD inhibited cell proliferation, reduced migration, repressed the cell cycle, and promoted cell apoptosis in GC cells. Based on RNA sequencing results and luciferase reporter assays, 3'-phosphoadenosine-5'-phosphosulfate synthase 2 (PAPSS2) was identified as a target gene of tRF-41-YDLBRY73W0K5KKOVD. These findings indicated that tRF-41-YDLBRY73W0K5KKOVD inhibited GC progression, suggesting that tRF-41-YDLBRY73W0K5KKOVD might be a potential therapeutic target in GC.

Identification of Metastasis-Associated Genes in Cutaneous Squamous Cell Carcinoma Based on Bioinformatics Analysis and Experimental Validation.

INTRODUCTION: Cutaneous squamous cell carcinoma (cSCC) is a global malignant tumor with a high degree of malignancy. Once metastasis occurs, it will lead to poor prognosis and even death. This study attempts to find out the central genes closely related to cSCC metastasis, so as to clarify the molecular regulatory mechanism of cSCC metastasis and open up new ideas for clinical treatment. METHODS: Firstly, cSCC data set GSE98767 was used to establish a tumor metastasis model via clustering analysis. The key module and hub genes associated with cSCC metastasis were analyzed by weighted gene co-expression analysis (WGCNA). Next, the prognostic functions of hub genes were identified by functional and pathway enrichment analysis, pan-cancer analysis, and receiver operating characteristic-area under the curve (ROC-AUC) validation. Finally, the key genes were verified by clinical sample detection and biological in vitro test. RESULTS: A total of 19 hub genes related to cSCC metastasis were identified. They were highly expressed in cSCC metastatic tissues and were mainly enriched in cellular material and energy metabolism pathways. Overall survival (OS) and disease-free survival (DFS) results from pan-cancer analysis showed that eight and six highly expressed genes, respectively, with PAPSS2 and SCG5 had highly reliable ROC-AUC validation values and were poor prognostic factors. Clinical and biological tests also confirmed the upregulation of PAPSS2 and SCG5 in cSCC. Deletion of PAPSS2 and SCG5 resulted in decreased viability, migration, and invasion of A-431 cells. CONCLUSION: PAPSS2 and SCG5 may be important factors for cSCC metastasis, and they are involved in the regulation of cSCC cell viability, migration, and invasion.

Novel Inactivating Homozygous PAPSS2 Mutation in Two Siblings With Disproportionate Short Stature.

Background/Objective: Variants in PAPSS2 (3'-phosphoadenosine 5'-phosphosulfate synthetase 2) present with varying degrees of brachyolmia (short trunk, platyspondyly, mild long-bone abnormalities). Our objective is to present the phenotype of male and female siblings with the same novel inactivating variant in PAPSS2. Case Report: A Jordanian female (case 1), born to consanguineous parents, was referred at 10 years of age for short stature (SS). She had a normal laboratory workup, including normal growth hormone stimulation testing. Spinal x-rays done for clinical scoliosis revealed platyspondyly. She attained an adult height of 143.5 cm (-3 SD). Years later, her brother (case 2) was referred at 21 months of age for SS. His laboratory workup and bone age were normal. His growth velocity declined at 6 years of age, but normal growth factors did not suggest growth hormone deficiency. When he returned during puberty, disproportionate body measurements were noted. A skeletal survey revealed platyspondyly, increasing suspicion of growth plate pathology. Exome sequencing in the family revealed a homozygous variant, p.His496Pro (H496P) in PAPSS2 (NM_004670.3:c.1487A>C). Both parents carried the same variant. Discussion: PAPSS2 assists with the sulfonation of dehydroepiandrosterone (DHEA) to DHEA sulfate and the sulfonation of proteoglycans in the cartilage, necessary for endochondral bone formation. PAPSS2-inactivating variants present with skeletal dysplasia and elevated DHEA levels. Conclusion: This novel variant in PAPSS2 manifested with mild brachyolmia but disproportionate SS in male and female siblings. Biochemical phenotype with low circulating DHEA sulfate and high DHEA levels reflect a sulfonation defect.

Inhibition of p53 Sulfoconjugation Prevents Oxidative Hepatotoxicity and Acute Liver Failure.

BACKGROUND & AIMS: Sulfoconjugation of small molecules or protein peptides is a key mechanism to ensure biochemical and functional homeostasis in mammals. The PAPS synthase 2 (PAPSS2) is the primary enzyme to synthesize the universal sulfonate donor 3'-phosphoadenosine 5'-phosphosulfate (PAPS). Acetaminophen (APAP) overdose is the leading cause of acute liver failure (ALF), in which oxidative stress is a key pathogenic event, whereas sulfation of APAP contributes to its detoxification. The goal of this study was to determine whether and how PAPSS2 plays a role in APAP-induced ALF. METHODS: Gene expression was analyzed in APAP-induced ALF in patients and mice. Liver-specific Papss2-knockout mice using Alb-Cre (Papss2ΔHC) or AAV8-TBG-Cre (Papss2iΔHC) were created and subjected to APAP-induced ALF. Primary human and mouse hepatocytes were used for in vitro mechanistic analysis. RESULTS: The hepatic expression of PAPSS2 was decreased in APAP-induced ALF in patients and mice. Surprisingly, Papss2ΔHC mice were protected from APAP-induced hepatotoxicity despite having a decreased APAP sulfation, which was accompanied by increased hepatic antioxidative capacity through the activation of the p53-p2-Nrf2 axis. Treatment with a sulfation inhibitor also ameliorated APAP-induced hepatotoxicity. Gene knockdown experiments showed that the hepatoprotective effect of Papss2ΔHC was Nrf2, p53, and p21 dependent. Mechanistically, we identified p53 as a novel substrate of sulfation. Papss2 ablation led to p53 protein accumulation by preventing p53 sulfation, which disrupts p53-MDM2 interaction and p53 ubiquitination and increases p53 protein stability. CONCLUSIONS: We have uncovered a previously unrecognized and p53-mediated role of PAPSS2 in controlling oxidative response. Inhibition of p53 sulfation may be explored for the clinical management of APAP overdose.

Intestinal Sulfation Is Essential to Protect Against Colitis and Colonic Carcinogenesis.

BACKGROUND & AIMS: Sulfation is a conjugation reaction essential for numerous biochemical and cellular functions in mammals. The 3'-phosphoadenosine 5'-phosphosulfate (PAPS) synthase 2 (PAPSS2) is the key enzyme to generate PAPS, which is the universal sulfonate donor for all sulfation reactions. The goal of this study was to determine whether and how PAPSS2 plays a role in colitis and colonic carcinogenesis. METHODS: Tissue arrays of human colon cancer specimens, gene expression data, and clinical features of cancer patients were analyzed. Intestinal-specific Papss2 knockout mice (Papss2ΔIE) were created and subjected to dextran sodium sulfate-induced colitis and colonic carcinogenesis induced by a combined treatment of azoxymethane and dextran sodium sulfate or azoxymethane alone. RESULTS: The expression of PAPSS2 is decreased in the colon cancers of mice and humans. The lower expression of PAPSS2 in colon cancer patients is correlated with worse survival. Papss2ΔIE mice showed heightened sensitivity to colitis and colon cancer by damaging the intestinal mucosal barrier, increasing intestinal permeability and bacteria infiltration, and worsening the intestinal tumor microenvironment. Mechanistically, the Papss2ΔIE mice exhibited reduced intestinal sulfomucin content. Metabolomic analyses revealed the accumulation of bile acids, including the Farnesoid X receptor antagonist bile acid tauro-ß-muricholic acid, and deficiency in the formation of bile acid sulfates in the colon of Papss2ΔIE mice. CONCLUSIONS: We have uncovered an important role of PAPSS2-mediated sulfation in colitis and colonic carcinogenesis. Intestinal sulfation may represent a potential diagnostic marker and PAPSS2 may serve as a potential therapeutic target for inflammatory bowel disease and colon cancer.

Steroid disulfates - Sulfation double trouble.

Sulfation pathways have recently come into the focus of biomedical research. For steroid hormones and related compounds, sulfation represents an additional layer of regulation as sulfated steroids are more water-soluble and tend to be biologically less active. For steroid diols, an additional sulfation is possible, carried out by the same sulfotransferases that catalyze the first sulfation step. The steroid disulfates that are formed are the focus of this review. We discuss both their biochemical production as well as their putative biological function. Steroid disulfates have also been linked to various clinical conditions in numerous untargeted metabolomics studies. New analytical techniques exploring the biosynthetic routes of steroid disulfates have led to novel insights, changing our understanding of sulfation in human biology. They promise a bright future for research into sulfation pathways, hopefully too for the diagnosis and treatment of several associated diseases.

Low DHEAS Concentration in a Girl Presenting with Short Stature and Premature Pubarche: A Novel PAPSS2 Gene Mutation.

OBJECTIVE: Dehydroepiandrosterone (DHEA) sulfotransferase (SULT2A1) converts DHEA to DHEA sulfate (DHEAS) which prevents bioactive androgen excess. This enzymatic reaction requires PAPS (3'-phospho-adenosine-5'-phosphosulfate) biosynthesis mediated by PAPS synthase 2 (PAPSS2). Here, we report a patient presenting with short stature and premature pubarche due to a novel homozygous mutation in the PAPPS2 gene. CASE REPORT: A 7.5-year-old girl was referred for short stature. She was born at term with a birth weight of 2,850 g and her parents were first cousins. At presentation, her height was 113.0 cm (-2.1 SDS) and weight was 28.3 kg (+0.9 SDS), her arm span was 115.0 cm, and upper to lower segment ratio was 1.2. Her pubic hair and breast development were at Tanner stage III and I, respectively. Radiographs revealed mild lumbar scoliosis and platyspondyly and irregular vertebral endplates in the thoracolumbar region. Her serum DHEAS was low (39 ng/mL). The plasma DHEAS/DHEA ratio was significantly decreased on 2 separate measurements (4.4 and 19.8; normal range 31-345). PAPSS2 gene analysis identified a homozygous p.L440Wfs*12 (c.1318_1330 delCTACTACACCCTC) variant. This is the first report of a large deletion leading to a frameshift effect in the PAPSS2 gene and a truncated PAPSS2 protein. CONCLUSION: We describe the third case with PAPSS2 deficiency presenting with premature pubarche, and the first large deletion in the PAPSS2 gene. Although PAPSS2 deficiency is a rare cause of premature pubarche and adrenal androgen excess, it should be considered, especially in cases with disproportionate short stature and clinical hyperandrogenism associated with low plasma DHEAS concentration.

PAPSS2-related brachyolmia: Clinical and radiological phenotype in 18 new cases.

Brachyolmia is a skeletal dysplasia characterized by short spine-short stature, platyspondyly, and minor long bone abnormalities. We describe 18 patients, from different ethnic backgrounds and ages ranging from infancy to 19 years, with the autosomal recessive form, associated with PAPSS2. The main clinical features include disproportionate short stature with short spine associated with variable symptoms of pain, stiffness, and spinal deformity. Eight patients presented prenatally with short femora, whereas later in childhood their short-spine phenotype emerged. We observed the same pattern of changing skeletal proportion in other patients. The radiological findings included platyspondyly, irregular end plates of the elongated vertebral bodies, narrow disc spaces and short over-faced pedicles. In the limbs, there was mild shortening of femoral necks and tibiae in some patients, whereas others had minor epiphyseal or metaphyseal changes. In all patients, exome and Sanger sequencing identified homozygous or compound heterozygous PAPSS2 variants, including c.809G>A, common to white European patients. Bi-parental inheritance was established where possible. Low serum DHEAS, but not overt androgen excess was identified. Our study indicates that autosomal recessive brachyolmia occurs across continents and may be under-recognized in infancy. This condition should be considered in the differential diagnosis of short femora presenting in the second trimester.

SOX9 protein is stabilized by TGF-ß and regulates PAPSS2 mRNA expression in chondrocytes.

OBJECTIVE: We previously identified 3'-phosphoadenosine 5'-phosphosulfate synthase 2 (PAPSS2) as a transcriptional target of transforming growth factor ß (TGF-ß) in chondrocytes. PAPSS2 is required for proper sulfation of proteoglycans in cartilage. Defective sulfation in the matrix results in alterations in mechanical properties of the cartilage that would be expected to result in degeneration. The objective of this study was to identify factors that regulate PAPSS2 expression and compare to a known TGF-ß responsive gene, proteoglycan 4/lubricin (PRG4). In this study, TGF-ß-mediated regulation of SOX9 was characterized, and the involvement of SOX9 in regulation of PAPSS2 mRNA was investigated. DESIGN: Primary bovine articular chondrocytes grown in micromass culture and ATDC5 cells were used as the model system. Adenoviruses were used to express SOX9 and SMAD3. siRNA was used to knock-down Sox9 and Smad3. Western blot and real-time quantitative RT-PCR (qPCR) were used to measure changes in protein and mRNA levels in response to treatment. RESULTS: Over-expression of SOX9 was sufficient to up-regulate PAPSS2 mRNA. TGF-ß treatment of SOX9-expressing cells resulted in enhanced up-regulation of PAPSS2 mRNA, suggesting that SOX9 cooperates with TGF-ß signaling. Furthermore, Sox9 was required for full TGF-ß-mediated induction of Papss2. In contrast, PRG4 was regulated by SMAD3 but not SOX9. SOX9 protein levels were increased after treatment with TGF-ß, although SOX9 mRNA was not. SOX9 protein was post-translationally stabilized after treatment with TGF-ß. CONCLUSIONS: TGF-ß stabilizes SOX9 protein, and SOX9 is sufficient and necessary for TGF-ß-mediated regulation of PAPSS2 mRNA, providing a novel mechanism for TGF-ß-mediated gene regulation in chondrocytes.

Characterizing drug-metabolizing enzymes and transporters that are bona fide CAR-target genes in mouse intestine.

Intestine is responsible for the biotransformation of many orally-exposed chemicals. The constitutive androstane receptor (CAR/Nr1i3) is known to up-regulate many genes encoding drug-metabolizing enzymes and transporters (drug-processing genes/DPGs) in liver, but less is known regarding its effect in intestine. Sixty-day-old wild-type and Car-/- mice were administered the CAR-ligand TCPOBOP or vehicle once daily for 4 days. In wild-type mice, Car mRNA was down-regulated by TCPOBOP in liver and duodenum. Car-/- mice had altered basal intestinal expression of many DPGs in a section-specific manner. Consistent with the liver data (Aleksunes and Klaassen, 2012), TCPOBOP up-regulated many DPGs (Cyp2b10, Cyp3a11, Aldh1a1, Aldh1a7, Gsta1, Gsta4, Gstm1-m4, Gstt1, Ugt1a1, Ugt2b34, Ugt2b36, and Mrp2-4) in specific sections of small intestine in a CAR-dependent manner. However, the mRNAs of Nqo1 and Papss2 were previously known to be up-regulated by TCPOBOP in liver but were not altered in intestine. Interestingly, many known CAR-target genes were highest expressed in colon where CAR is minimally expressed, suggesting that additional regulators are involved in regulating their expression. In conclusion, CAR regulates the basal expression of many DPGs in intestine, and although many hepatic CAR-targeted DPGs were bona fide CAR-targets in intestine, pharmacological activation of CAR in liver and intestine are not identical.

Autosomal recessive brachyolmia: early radiological findings.

Brachyolmia (BO) is a heterogeneous group of skeletal dysplasias with skeletal changes limited to the spine or with minimal extraspinal features. BO is currently classified into types 1, 2, 3, and 4. BO types 1 and 4 are autosomal recessive conditions caused by PAPSS2 mutations, which may be merged together as an autosomal recessive BO (AR-BO). The clinical and radiological signs of AR-BO in late childhood have already been reported; however, the early manifestations and their age-dependent evolution have not been well documented. We report an affected boy with AR-BO, whose skeletal abnormalities were detected in utero and who was followed until 10 years of age. Prenatal ultrasound showed bowing of the legs. In infancy, radiographs showed moderate platyspondyly and dumbbell deformity of the tubular bones. Gradually, the platyspondyly became more pronounced, while the bowing of the legs and dumbbell deformities of the tubular bones diminished with age. In late childhood, the overall findings were consistent with known features of AR-BO. Genetic testing confirmed the diagnosis. Being aware of the initial skeletal changes may facilitate early diagnosis of PAPSS2-related skeletal dysplasias.

Gene Expression Differences Between Offspring of Long-Lived Individuals and Controls in Candidate Longevity Regions: Evidence for PAPSS2 as a Longevity Gene.

Although there is compelling evidence for a genetic contribution to longevity, identification of specific genes that robustly associate with longevity has been a challenge. In order to identify longevity-enhancing genes, we measured differential gene expression between offspring of long-lived Amish (older than 90 years; cases, n = 128) and spouses of these offspring (controls, n = 121) and correlated differentially expressed transcripts with locations of longevity-associated variants detected in a prior genome-wide association study (GWAS) of survival to age 90. Expression of one of these transcripts, 3'-phosphoadenosine 5'-phosphosulfate synthase 2 (PAPSS2), was significantly higher in offspring versus controls (4×10(-4)) and this association was replicated using quantitative real-time polymerase chain reaction. PAPSS2, a sulfation enzyme located on chromosome 10, is ~80kb upstream of the PAPSS2 transcription start site. We found evidence of cis-expression for the originally reported GWAS SNP and PAPSS2 Monogenic conditions linked to PAPSS2 include andrenocortical androgen excess resulting in premature pubarche and skeletal dysplasias, both of which have premature aging features. In summary, these findings provide novel evidence for PAPSS2 as a longevity locus and illustrate the value of harnessing multiple "-omic" approaches to identify longevity candidates.

Clinical and radiographic features of the autosomal recessive form of brachyolmia caused by PAPSS2 mutations.

Brachyolmia is a heterogeneous skeletal dysplasia characterized by generalized platyspondyly without significant long-bone abnormalities. Based on the mode of inheritance and radiographic features, at least three types of brachyolmia have been postulated. We recently identified an autosomal recessive form of brachyolmia that is caused by loss-of-function mutations of PAPSS2, the gene encoding PAPS (3'-phosphoadenosine 5'-phosphosulfate) synthase 2. To understand brachyolmia caused by PAPSS2 mutations (PAPSS2-brachyolmia), we extended our PAPSS2 mutation analysis to 13 patients from 10 families and identified homozygous or compound heterozygous mutations in all. Nine different mutations were found: three splice donor-site mutations, three missense mutations, and three insertion or deletion mutations within coding regions. In vitro enzyme assays showed that the missense mutations were also loss-of-function mutations. Phenotypic characteristics of PAPSS2-brachyolmia include short-trunk short stature, normal intelligence and facies, spinal deformity, and broad proximal interphalangeal joints. Radiographic features include platyspondyly with rectangular vertebral bodies and irregular end plates, broad ilia, metaphyseal changes of the proximal femur, including short femoral neck and striation, and dysplasia of the short tubular bones. PAPSS2-brachyolmia includes phenotypes of the conventional clinical concept of brachyolmia, the Hobaek and Toledo types, and is associated with abnormal androgen metabolism.