CUBN gene mutations may cause focal segmental glomerulosclerosis (FSGS) in children

Background Imerslund-Gräsbeck Syndrome (IGS) is mainly caused by CUBN gene biallelic mutations. Proteinuria accompanies IGS specific symptoms in about half of the patients, isolated proteinuria is rarely reported. Here we present 3 patients with isolated proteinuria and focal segmental glomerulosclerosis (FSGS) caused by CUBN gene biallelic pathogenic variants. Method Whole exome sequencing was performed on three children with isolated proteinuria. CUBN gene biallelic pathogenic variants were found and then verified by sanger sequencing. Their clinical, pathological and molecular genetic characteristics were analyzed and correlated accordingly. Results All three children presented with isolated proteinuria, no megaloblastic anemia. Their urine levels of β2 microglobulin were normal or slightly higher. Renal biopsies showed focal segmental glomerulosclerosis with mild glomerular mesangial hypercellularity, partial effacement of foot processes and podocyte microvillation. Two of them were found to carry compound heterozygous mutations and one homozygous mutation of CUBN gene. Totally four CUBN gene biallelic pathogenic variants were identified, including c.9287 T > C (p.L3096P), c.122 + 1G > A, c.7906C > T (p.R2636*), c.10233G > A (p.W3411*). Except for intron splice-site mutation, all other variants are located in highly conserved sites of CUB domain for binding to albumin. Conclusion The results demonstrate that CUBN gene mutations may cause isolated proteinuria pathologically presented as FSGS. Our cases extend the spectrum of renal manifestation and genotype of CUBN gene mutations.

Splice Junction Identification using Long Short-Term Memory Neural Networks

Background: Splice junctions are the key to going from pre-messenger RNA to mature messenger RNA in many multi-exon genes due to alternative splicing. Since the percentage of multi-exon genes that undergo alternative splicing is very high, identifying splice junctions is an attractive research topic with important implications. Objective: The aim is to develop a deep learning model capable of identifying splice junctions in RNA sequences using 13,666 unique sequences of primate RNA. Method: A Long Short-Term Memory (LSTM) Neural Network model is developed that classifies a given sequence as EI (Exon-Intron splice), IE (Intron-Exon splice), or N (No splice). The model is trained with groups of trinucleotides and its performance is tested using validation and test data to prevent bias. Results: Model performance was measured using accuracy and f-score in test data. The finalized model achieved an average accuracy of 91.34% with an average f-score of 91.36% over 50 runs. Conclusion: Comparisons show a highly competitive model to recent Convolutional Neural Network structures. The proposed LSTM model achieves the highest accuracy and f-score among published alternative LSTM structures.

CUBN Gene Mutations May Cause Focal Segmental Glomerulosclerosis (FSGS) in Children

Background: CUBN gene mutation is extremely rare and thought to only presented as tubular proteinuria without glomerular involvement. Here, we present 3 patients with prominent proteinuria and FSGS in renal pathologies caused by novel CUBN gene mutations.Method: Whole exome sequencing was performed in three children. CUBN gene mutations were found and then verified by sanger sequencing. Their clinical, pathological and molecular genetic characteristics were analyzed and correlated accordingly. Results: All three children presented with prominent proteinuria but normal or slightly higher levels of urine β2 microglobulin, without megaloblastic anemia. Renal biopsies showed segmental glomerular sclerosis, glomerular mesangial cells proliferation, effacement of foot processes, podocyte microvillation and interstitial fibrosis. There were four novel CUBN gene mutations in our study, including c.9287T>C (p.L3096P), c.122+1G>A, c.7906C>T (p.R2636*), c.10233G>A (p.W3411*). Except for intron splice-site mutation, all other variants are located in highly conserved sites of CUB domain. Conclusion: In this study, four novel pathogenic mutations of CUBN gene were identified in three prominent proteinuric children. The results demonstrate that CUBN gene mutations may cause pathological changes of podocytes and FSGS, which was not previously reported. Our cases extend the spectrum of renal manifestation and genotype of CUBN gene mutation.

Genetic Diversity Analysis of Salvadora Oleoides Decne: An Evergreen Dry Land Fruit Tree of Rajasthan, India

Genetic diversity of Salvadora oleoides Decne is analyzed by cumulative data of 10 Random Amplified Polymorphic DNA (RAPD), 10 Inter Simple Sequence Repeats (ISSR) and 7 Intron Splice Junction (ISJ) markers. The plant is an evergreen fruit tree and well distributed in semi-arid and sub-humid climatic conditions of Rajasthan, India. RAPD, ISSR and ISJ primers accounted for 84.4%, 85.3%, 85.9% polymorphism. Average 0.23 PIC is accounted for RAPD, ISSR and ISJ primers. The genetic similarity ranged between 0.42-0.89. Analysis of molecular variance (AMOVA) revealed higher variation (73%) at intra-population than inter-population (27%) level. Genetic distances based on Un-weighted Pair Group Method with Arithmetic mean (UPGMA) dendrogram and Principal Coordinate Analysis (PCoA) is correlated with physical distances or climatic conditions of Salvadora oleoides Decne in a semi-arid and sub-humid environment of Rajasthan. The present investigation may help in the understanding of gene flow systems between physical distances and environmental heterogeneity of the populations for better management of Salvadora oleoides Decne in the region.

A mutation in the intron splice acceptor site of a GA3ox gene confers dwarf architecture in watermelon (Citrullus lanatus L.)

Dwarf architecture is an important trait associated with plant yield, lodging resistance and labor cost. Here, we aimed to identify a gene causing dwarfism in watermelon. The ‘w106’ (dwarf) and ‘Charleston Gray’ (vine) were used as parents to construct F1 and F2 progeny. Dwarf architecture of ‘w106’ was mainly caused by longitudinal cell length reduction and was controlled by a single recessive gene. Whole-genome sequencing of two parents and two bulk DNAs of F2 population localized this gene to a 2.63-Mb region on chromosome 9; this was further narrowed to a 541-kb region. Within this region, Cla015407, encoding a gibberellin 3β-hydroxylase (GA3ox), was the candidate gene. Cla015407 had a SNP mutation (G → A) in the splice acceptor site of the intron, leading to altered splicing event and generating two splicing isoforms in dwarf plants. One splicing isoform retained the intron sequences, while the other had a 13-bp deletion in the second exon of GA3ox transcript, both resulting in truncated proteins and loss of the functional Fe2OG dioxygenase domain in dwarf plants. RNA-Seq analysis indicated that expression of Cla015407 and other GA biosynthetic and metabolic genes were mostly up-regulated in the shoots of dwarf plants compared with vine plants in F2 population. Measurement of endogenous GA levels indicated that bioactive GA4 was significantly decreased in the shoots of dwarf plants. Moreover, the dwarf phenotype can be rescued by exogenous applications of GA3 or GA4+7, with the latter having a more distinct effect than the former. Subcellular localization analyses of GA3ox proteins from two parents revealed their subcellular targeting in nucleus and cytosol. Here, a GA3ox gene controlling dwarf architecture was identified, and loss function of GA3ox leads to GA4 reduction and dwarfism phenotype in watermelon.

A Silent Exonic Mutation in a Rice Integrin-α FG-GAP Repeat-Containing Gene Causes Male-Sterility by Affecting mRNA Splicing

Pollen development plays crucial roles in the life cycle of higher plants. Here we characterized a rice mutant with complete male-sterile phenotype, pollen-less 1 (pl1). pl1 exhibited smaller anthers with arrested pollen development, absent Ubisch bodies, necrosis-like tapetal hypertrophy, and smooth anther cuticular surface. Molecular mapping revealed a synonymous mutation in the fourth exon of PL1 co-segregated with the mutant phenotype. This mutation disrupts the exon-intron splice junction in PL1, generating aberrant mRNA species and truncated proteins. PL1 is highly expressed in the tapetal cells of developing anther, and its protein is co-localized with plasma membrane (PM) and endoplasmic reticulum (ER) signal. PL1 encodes an integrin-α FG-GAP repeat-containing protein, which has seven β-sheets and putative Ca2+-binding motifs and is broadly conserved in terrestrial plants. Our findings therefore provide insights into both the role of integrin-α FG-GAP repeat-containing protein in rice male fertility and the influence of exonic mutation on intronic splice donor site selection.

Potential G-quadruplex forming sequences and N6-methyladenosine colocalize at human pre-mRNA intron splice sites

ABSTRACTUsing bioinformatic analysis of published data, we identify in human mRNA that potential G-quadruplex forming sequences (PQSs) colocalize with the epitranscriptomic modifications N6-methyladenosine (m6A), pseudouridine (Ψ), and inosine (I). A deeper analysis of the colocalized m6A and PQSs found them intronic in pre-mRNA near 5′ and 3′ splice sites. The loop lengths and sequence context of the m6A-bearing PQSs found short loops most commonly comprised of A nucleotides. This observation is consistent with literature reports of intronic m6A found in SAG (S = C or G) consensus motifs that are also recognized by splicing factors. The localization of m6A and PQSs in pre-mRNA at intron splice junctions suggests that these features could be involved in alternative mRNA splicing. A similar analysis for PQSs around sites of Ψ installation or A-to-I editing in mRNA also found a colocalization; however, the frequency was less than that observed with m6A.TOC Graphic