RFX2 is a candidate downstream amplifier of A-MYB regulation in mouse spermatogenesis

<p>Abstract</p> <p>Background</p> <p>Mammalian spermatogenesis involves formation of haploid cells from the male germline and then a complex morphological transformation to generate motile sperm. Focusing on meiotic prophase, some tissue-specific transcription factors are known (A-MYB) or suspected (RFX2) to play important roles in modulating gene expression in pachytene spermatocytes. The current work was initiated to identify both downstream and upstream regulatory connections for <it>Rfx2</it>.</p> <p>Results</p> <p>Searches of pachytene up-regulated genes identified high affinity RFX binding sites (X boxes) in promoter regions of several new genes: <it>Adam5</it>, <it>Pdcl2</it>, and <it>Spag6</it>. We confirmed a strong promoter-region X-box for <it>Alf</it>, a germ cell-specific variant of general transcription factor TFIIA. Using <it>Alf </it>as an example of a target gene, we showed that its promoter is stimulated by RFX2 in transfected cells and used ChIP analysis to show that the promoter is occupied by RFX2 in vivo. Turning to upstream regulation of the <it>Rfx2 </it>promoter, we identified a cluster of three binding sites (MBS) for the MYB family of transcription factors. Because testis is one of the few sites of <it>A-myb </it>expression, and because spermatogenesis arrests in pachytene in <it>A-myb </it>knockout mice, the MBS cluster implicates <it>Rfx2 </it>as an <it>A-myb </it>target. Electrophoretic gel-shift, ChIP, and co-transfection assays all support a role for these MYB sites in <it>Rfx2 </it>expression. Further, <it>Rfx2 </it>expression was virtually eliminated in <it>A-myb </it>knockout testes. Immunohistology on testis sections showed that A-MYB expression is up-regulated only after pachytene spermatocytes have clearly moved away from the tubule wall, which correlates with onset of RFX2 expression, whereas B-MYB expression, by contrast, is prevalent only in earlier spermatocytes and spermatogonia.</p> <p>Conclusion</p> <p>With an expanding list of likely target genes, RFX2 is potentially an important transcriptional regulator in pachytene spermatocytes. <it>Rfx2 </it>itself is a good candidate to be regulated by A-MYB, which is essential for meiotic progression. If <it>Alf </it>is a genuine RFX2 target, then <it>A-myb</it>, <it>Rfx2</it>, and <it>Alf </it>may form part of a transcriptional network that is vital for completion of meiosis and preparation for post-meiotic differentiation.</p

Ultrastructural immunolocalization of histones (H2B, H3, H4), transition protein (TP1) and protamine in rabbit spermatids and spermatozoa nuclei. Relation to condensation of the chromatin

The histones H2B, H3 and H4, the transition protein TP1 and protamine were localised using ultrastructural immunocytochemistry in nuclei of rabbit spermatids and spermatozoa. Histones are present in round spermatid nuclei and are lost during the elongation of nuclei. TP1 and protamine appear simultaneously in all nuclei during this period. TP1 is located at the periphery of chromatin cords, while protamine seems to be located at random in the same cords. TP1 is lost in most elongated sprematids during step 13 of spermiogenesis, and the protamine stays in all sperm nuclei. TP1 remains persent in some old spermatids and ejaculated spermatozoa. In the rabbit, 3--6% of sperm nuclei decondense spontaneously. Most are characterized by a retention of TP1. Respective roles of TP1 and the protamine in spermatid nuclear condensation are discussed

A Cytochemical Study of the Transcriptional and Translational Regulation of Nuclear Transition Protein 1 (TP1), a Major Chromosomal Protein of Mammalian Spermatids

Immunocytochemical localization and in situ hybridization techniques were used to investigate the presence of spermatid nuclear transition protein 1 (TP1) and its mRNA during the various stages of spermatogenesis in the rat. A specific antiserum to TP1 was raised in a rabbit and used to show that TP1 is immunologically crossreactive among many mammals including humans. During spermatogenesis the protein appears in spermatids as they progress from step 12 to step 13, a period in which nuclear condensation is underway. The protein is lost during step 15. An asymmetric RNA probe generated from a TP1 cDNA clone identified TP1 mRNA in late round spermatids beginning in step 7. The message could no longer be detected in spermatids of step 15 or beyond. Thus, TP1 mRNA first appears well after meiosis in haploid cells but is not translated effectively for the several days required for these cells to progress to the stage of chromatin condensation. Message and then protein disappear as the spermatids enter step 15. In agreement with a companion biochemical study (Heidaran, M.A., and W.S. Kistler. J. Biol. Chem. 1987. 262:13309-13315), these results establish that translational control is involved in synthesis of this major spermatid nuclear protein. In addition, they suggest that TP1 plays a role in the completion but not the initiation of chromatin condensation in elongated spermatids

Description of understory development in a tree plantation with a new method of data structuring

During May and June, 1974, relevés were obtained from 30 plantation stands in the Saginaw Forest in southeastern Michigan. The canopy trees in these plantations were planted between 1904 and 1938. The understory has developed naturally over the years. The forest plantations offer opportunities for study of the effects of the canopy on the structure and species composition of the understory.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/43888/1/11258_2004_Article_BF00228485.pd

Precision Departure Release Capability (PDRC) Final Report

After takeoff, aircraft must merge into en route (Center) airspace traffic flows that may be subject to constraints that create localized demand/capacity imbalances. When demand exceeds capacity, Traffic Management Coordinators (TMCs) and Frontline Managers (FLMs) often use tactical departure scheduling to manage the flow of departures into the constrained Center traffic flow. Tactical departure scheduling usually involves a Call for Release (CFR) procedure wherein the Tower must call the Center to coordinate a release time prior to allowing the flight to depart. In present-day operations release times are computed by the Center Traffic Management Advisor (TMA) decision support tool, based upon manual estimates of aircraft ready time verbally communicated from the Tower to the Center. The TMA-computed release time is verbally communicated from the Center back to the Tower where it is relayed to the Local controller as a release window that is typically three minutes wide. The Local controller will manage the departure to meet the coordinated release time window. Manual ready time prediction and verbal release time coordination are labor intensive and prone to inaccuracy. Also, use of release time windows adds uncertainty to the tactical departure process. Analysis of more than one million flights from January 2011 indicates that a significant number of tactically scheduled aircraft missed their en route slot due to ready time prediction uncertainty. Uncertainty in ready time estimates may result in missed opportunities to merge into constrained en route flows and lead to lost throughput. Next Generation Air Transportation System plans call for development of Tower automation systems capable of computing surface trajectory-based ready time estimates. NASA has developed the Precision Departure Release Capability (PDRC) concept that improves tactical departure scheduling by automatically communicating surface trajectory-based ready time predictions and departure runway assignments to the Center scheduling tool. The PDRC concept also incorporates earlier NASA and FAA research into automation-assisted CFR coordination. The PDRC concept reduces uncertainty by automatically communicating coordinated release times with seconds-level precision enabling TMCs and FLMs to work with target times rather than windows. NASA has developed a PDRC prototype system that integrates the Center's TMA system with a research prototype Tower decision support tool. A two-phase field evaluation was conducted at NASA's North Texas Research Station in Dallas/Fort Worth. The field evaluation validated the PDRC concept and demonstrated reduced release time uncertainty while being used for tactical departure scheduling of more than 230 operational flights over 29 weeks of operations. This paper presents research results from the PDRC research activity. Companion papers present the Concept of Operations and a Technology Description

Asymptotic and numerical analysis of a simple model for blade coating

Motivated by the industrial process of blade coating, the two-dimensional flow of a thin film of Newtonian fluid on a horizontal substrate moving parallel to itself with constant speed under a fixed blade of finite length in which the flows upstream and downstream of the blade are coupled via the flow under the blade is analysed. A combination of asymptotic and numerical methods is used to investigate the number and nature of the steady solutions that exist. Specially, it is found that in the presence of gravity there is always at least one, and (depending on the parameter values) possibly as many as three, steady solutions, and that when multiple solutions occur they are identical under and downstream of the blade, but differ upstream of it. The stability of these solutions is investigated, and their asymptotic behaviour in the limits of large and small flux and weak and strong gravity effects, respectively, determined

Transplantation for renal failure secondary to enteric hyperoxaluria: a case report

Enteric hyperoxaluria can lead to renal failure. There have only been a few reports of renal transplantation as treatment of endstage renal disease secondary to enteric hyperoxaluria and results have been mixed. This report describes a patient with Crohn's disease who developed chronic renal failure from enteric hyperoxaluria. He subsequently had a successful renal transplant without any post-operative oxalate related complications and has satisfactory renal function almost three years later. Aggressive pre-transplant hemodialysis was not done. The literature associated with renal transplantation for enteric hyperoxaluria is reviewed

Interplay of RFX transcription factors 1, 2 and 3 in motile ciliogenesis

Cilia assembly is under strict transcriptional control during animal development. In vertebrates, a hierarchy of transcription factors (TFs) are involved in controlling the specification, differentiation and function of multiciliated epithelia. RFX TFs play key functions in the control of ciliogenesis in animals. Whereas only one RFX factor regulates ciliogenesis in C. elegans, several distinct RFX factors have been implicated in this process in vertebrates. However, a clear understanding of the specific and redundant functions of different RFX factors in ciliated cells remains lacking. Using RNA-seq and ChIP-seq approaches we identified genes regulated directly and indirectly by RFX1, RFX2 and RFX3 in mouse ependymal cells. We show that these three TFs have both redundant and specific functions in ependymal cells. Whereas RFX1, RFX2 and RFX3 occupy many shared genomic loci, only RFX2 and RFX3 play a prominent and redundant function in the control of motile ciliogenesis in mice. Our results provide a valuable list of candidate ciliary genes. They also reveal stunning differences between compensatory processes operating in vivo and ex vivo

Mass‐loading the Earth's dayside magnetopause boundary layer and its effect on magnetic reconnection

When the interplanetary magnetic field is northward for a period of time, O+ from the high‐latitude ionosphere escapes along reconnected magnetic field lines into the dayside magnetopause boundary layer. Dual‐lobe reconnection closes these field lines, which traps O+ and mass loads the boundary layer. This O+ is an additional source of magnetospheric plasma that interacts with magnetosheath plasma through magnetic reconnection. This mass loading and interaction is illustrated through analysis of a magnetopause crossing by the Magnetospheric Multiscale spacecraft. While in the O+‐rich boundary layer, the interplanetary magnetic field turns southward. As the Magnetospheric Multiscale spacecraft cross the high‐shear magnetopause, reconnection signatures are observed. While the reconnection rate is likely reduced by the mass loading, reconnection is not suppressed at the magnetopause. The high‐latitude dayside ionosphere is therefore a source of magnetospheric ions that contributes often to transient reduction in the reconnection rate at the dayside magnetopause.publishedVersio

Distinguishing Molecular Features and Clinical Characteristics of a Putative New Rhinovirus Species, Human Rhinovirus C (HRV C)

Background: Human rhinoviruses (HRVs) are the most frequently detected pathogens in acute respiratory tract infections (ARTIs) and yet little is known about the prevalence, recurrence, structure and clinical impact of individual members. During 2007, the complete coding sequences of six previously unknown and highly divergent HRV strains were reported. To catalogue the molecular and clinical features distinguishing the divergent HRV strains, we undertook, for the first time, in silico analyses of all available polyprotein sequences and performed retrospective reviews of the medical records of cases in which variants of the prototype strain, HRV-QPM, had been detected